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Sample records for metaphase

  1. Amphitelic orientation of centromeres at metaphase I is an important ...

    Indian Academy of Sciences (India)

    2014-07-31

    Jul 31, 2014 ... Centromere orientation in meiotic metaphase I. Figure 1. Meiotic observation of hybrids between synthetic hexaploid wheat line Syn-SAU-6 and rye. Rye centromeres were detected with the probe PrCEN-1 (green). a, prophase; b, early metaphase; c, metaphase; d & e, late metaphase; f & g, anaphase; h, ...

  2. Dense chromatin plates in metaphase chromosomes.

    Science.gov (United States)

    Gállego, Isaac; Castro-Hartmann, Pablo; Caravaca, Juan Manuel; Caño, Silvia; Daban, Joan-Ramon

    2009-04-01

    In a previous work we observed multilayered plate-like structures surrounding partially denatured HeLa chromosomes at metaphase ionic conditions. This unexpected finding has led us to carry out an extensive investigation of these structures. Our results show that plates can also be found in metaphase chromosomes from chicken lymphocytes. We have used atomic force microscopy (AFM) to image and investigate the mechanical properties of plates in aqueous solution. Plates are thin (approximately 6.5 nm each layer) but compact and resistant to penetration by the AFM tip: their Young's modulus is approximately 0.2 GPa and the stress required for surface penetration is approximately 0.03 GPa in the presence of Mg(2+) (5-20 mM). Low-ionic strength conditions produce emanation of chromatin fibers from the edges of uncrosslinked plates. These observations and AFM results obtained applying high forces indicate that the chromatin filament is tightly tethered inside the plates. Images of metal-shadowed plates and cryo-electron microscopy images of frozen-hydrated plates suggest that nucleosomes are tilted with respect to the plate surface to allow an interdigitation between the successive layers and a thickness reduction compatible with the observed plate height. The similarities between denatured plates from chicken chromosomes and aggregates of purified chromatin from chicken erythrocytes suggest that chromatin has intrinsic structural properties leading to plate formation. Scanning electron micrographs and images obtained with the 200-kV transmission microscope show that plates are the dominant component of compact chromatids. We propose that metaphase chromosomes are formed by many stacked plates perpendicular to the chromatid axis.

  3. FISHprep: A Novel Integrated Device for Metaphase FISH Sample Preparation

    DEFF Research Database (Denmark)

    Shah, Pranjul Jaykumar; Vedarethinam, Indumathi; Kwasny, Dorota

    2011-01-01

    We present a novel integrated device for preparing metaphase chromosomes spread slides (FISHprep). The quality of cytogenetic analysis from patient samples greatly relies on the efficiency of sample pre-treatment and/or slide preparation. In cytogenetic slide preparation, cell cultures...... are routinely used to process samples (for culture, arrest and fixation of cells) and/or to expand limited amount of samples (in case of prenatal diagnostics). Arguably, this expansion and other sample pretreatments form the longest part of the entire diagnostic protocols spanning over 3–4 days. We present here...... with minimal handling for metaphase FISH slide preparation....

  4. Metaphase chromosome analysis by ligation-mediated PCR: heritable chromatin structure and a comparison of active and inactive X chromosomes.

    OpenAIRE

    Hershkovitz, M; Riggs, A D

    1995-01-01

    We report that ligation-mediated PCR (LMPCR) can be used for high-resolution study of metaphase chromosomes, and we discuss the role of metaphase chromatin structure in the preservation of differentiated cell states. The X chromosome-linked human PGK1 (phosphoglycerate kinase 1) promoter region was investigated, and euchromatic active X chromosome (Xa) metaphase chromatin was compared with interphase Xa chromatin and to heterochromatic inactive X chromosome (Xi) metaphase and interphase chrom...

  5. Interchromatidal central ridge and transversal symmetry in early metaphasic human chromosome one.

    Science.gov (United States)

    Argüello-Miranda, Orlando; Sáenz-Arce, Giovanni

    2008-01-01

    The topographic structure of Giemsa-banded (G-banded) early metaphase human chromosomes adsorbed on glass was analyzed by atomic force microscope using amplitude modulation mode (AM-AFM). Longitudinal height measurements for early metaphasic human chromosomes showed a central ridge that was further characterized by transversal height measurements. The heterochromatic regions displayed a high level of transversal symmetry, while the euchromatic ones presented several peaks across the transversal height measurements. We suggest that this central ridge and symmetry patterns point out a transitional arrangement of the early metaphase chromosome and support evidence for interchromatidal interactions prior to disjunction. 2008 John Wiley & Sons, Ltd

  6. A cost-effective and flexible metaphase finder system for chromosomal biodosimetry

    International Nuclear Information System (INIS)

    Furukawa, A.; Hayata, I.

    2003-01-01

    For biological dosimetry by counting chromosomes aberrations, automation technique has been required to process large number of sample preparations at low dose radiation exposure. Metaphase Finder is an automated optical microscope system, which automatically scans and finds metaphase cells on the slide glass, and relocates metaphase cells to the center of the field of view of the microscope to observe chromosomes in high magnification. There are several commercial products of metaphase finders, but these dedicated-system products are usually expensive and inconvenient for fitting to the variety of sample preparations. Now, we have constructed an improved system by assembling each components of the system, such as microscope, automated stage and computer, from the selection of the commercially available products, instead of purchasing one dedicated system. The new system has high cost-effectiveness and high flexibility in adapting to the new staining methods. This Metaphase Finder system consists of following components: an optical microscope, a motorized sample stage, an auto-focusing unit and two CCD cameras for focusing and for image acquisition. These components are controlled by a personal computer. The image recognition software for detecting metaphase cells from the microscope image was developed on the programming environment provided by the general-purpose image analysis software. The processing speed of this Metaphase Finder system is 20 to 30 minutes per one slide glass (typical scan area is 20 mm x 50 mm), which is enough acceptable for practical use and remarkably improved from the speed of our previous model of 1993. The algorithm for detecting metaphase cells has also improved to adapt to the variety of the condition of the sample preparation. This new system is scheduled to be distributed to several laboratories in Japan, and will be tested for the practical use

  7. Antagonistic spindle motors and MAPs regulate metaphase spindle length and chromosome segregation.

    Science.gov (United States)

    Syrovatkina, Viktoriya; Fu, Chuanhai; Tran, Phong T

    2013-12-02

    Metaphase describes a phase of mitosis where chromosomes are attached and oriented on the bipolar spindle for subsequent segregation at anaphase. In diverse cell types, the metaphase spindle is maintained at characteristic constant length [1-3]. Metaphase spindle length is proposed to be regulated by a balance of pushing and pulling forces generated by distinct sets of spindle microtubules (MTs) and their interactions with motors and MT-associated proteins (MAPs). Spindle length is further proposed to be important for chromosome segregation fidelity, as cells with shorter- or longer-than-normal metaphase spindles, generated through deletion or inhibition of individual mitotic motors or MAPs, showed chromosome segregation defects. To test the force-balance model of spindle length control and its effect on chromosome segregation, we applied fast microfluidic temperature control with live-cell imaging to monitor the effect of deleting or switching off different combinations of antagonistic force contributors in the fission yeast metaphase spindle. We show that the spindle midzone proteins kinesin-5 cut7p and MT bundler ase1p contribute to outward-pushing forces and that the spindle kinetochore proteins kinesin-8 klp5/6p and dam1p contribute to inward-pulling forces. Removing these proteins individually led to aberrant metaphase spindle length and chromosome segregation defects. Removing these proteins in antagonistic combination rescued the defective spindle length and in some combinations also partially rescued chromosome segregation defects. Copyright © 2013 Elsevier Ltd. All rights reserved.

  8. FahamecV1:A Low Cost Automated Metaphase Detection System

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    H. Yilmaz

    2017-12-01

    Full Text Available In this study, FahamecV1 is introduced and investigated as a low cost and high accuracy solution for metaphase detection. Chromosome analysis is performed at the metaphase stage and high accuracy and automated detection of the metaphase stage plays an active role in decreasing analysis time. FahamecV1 includes an optic microscope, a motorized microscope stage, an electronic control unit, a camera, a computer and a software application. Printing components of the motorized microscope stage (using a 3D printer is of the main reasons for cost reduction. Operations such as stepper motor calibration, are detection, focusing, scanning, metaphase detection and saving of coordinates into a database are automatically performed. To detect metaphases, a filter named Metafilter is developed and applied. Average scanning time per preparate is 77 sec/cm2. True positive rate is calculated as 95.1%, true negative rate is calculated as 99.0% and accuracy is calculated as 98.8%.

  9. AFM picking-up manipulation of the metaphase chromosome fragment by using the tweezers-type probe

    International Nuclear Information System (INIS)

    Yamanaka, Keiichiro; Saito, Masato; Shichiri, Motoharu; Sugiyama, Sigeru; Takamura, Yuzuru; Hashiguchi, Gen; Tamiya, Eiichi

    2008-01-01

    We have studied the development of a new procedure based on atomic force microscopy (AFM) for the analysis of metaphase chromosome. The aim of this study was to obtain detailed information about the specific locations of genes on the metaphase chromosome. In this research, we performed the manipulation of the metaphase chromosome by using novel AFM probes to obtain chromosome fragments of a smaller size than the ones obtained using the conventional methods, such as glass microneedles. We could pick up the fragment of the metaphase chromosome dissected by the knife-edged probe by using our tweezers-type probe

  10. Prognostic impact of a suboptimal number of analyzed metaphases in normal karyotype lower-risk MDS.

    Science.gov (United States)

    de Swart, Louise; Smith, Alex; Haase, Detlef; Fenaux, Pierre; Symeonidis, Argiris; Cermak, Jaroslav; Sanz, Guillermo; Stauder, Reinhard; Mittelman, Moshe; Hellström-Lindberg, Eva; Malcovati, Luca; Langemeijer, Saskia; Skov-Holm, Mette; Mądry, Krzysztof; Germing, Ulrich; Almeida, Antonio Medina; Tatic, Aurelia; Savic, Aleksandar; Šimec, Njetočka Gredelj; van Marrewijk, Corine; Guerci-Bresler, Agnes; Sanhes, Laurence; Luño, Elisa; Culligan, Dominic; Beyne-Rauzy, Odile; Burgstaller, Sonja; Blijlevens, Nicole; Bowen, David; de Witte, Theo

    2018-04-01

    Conventional karyotype is one of the most relevant prognostic factors in MDS. However, about 50% of patients with MDS have a normal karyotype. Usually, 20-25 normal metaphases (nMP) are considered to be optimal to exclude small abnormal clones which might be associated with poor prognosis. This study evaluated the impact of examining a suboptimal number of metaphases in patients recruited to the EUMDS Registry with low and intermediate-1 risk according to IPSS. Only 179/1049 (17%) of patients with a normal karyotype had a suboptimal number of nMP, defined as less than 20 metaphases analyzed. The outcome (overall survival and progression-free survival) of patients with suboptimal nMP was not inferior to those with higher numbers of analyzed MP both in univariate and multivariate analyses. For patients with an abnormal karyotype, 224/649 (35%) had a suboptimal number of MP assessed, but this did not impact on outcome. For patients with a normal karyotype and suboptimal numbers of analyzable metaphases standard evaluation might be acceptable for general practice, but we recommend additional FISH-analyses or molecular techniques, especially in candidates for intensive interventions. Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.

  11. Metaphase FISH on a Chip: Miniaturized Microfluidic Device for Fluorescence in situ Hybridization

    DEFF Research Database (Denmark)

    Vedarethinam, Indumathi; Shah, Pranjul Jaykumar; Dimaki, Maria

    2010-01-01

    -FISH, the process continues to be a manual, labour intensive, expensive and time consuming technique, often taking over 3-5 days, even in dedicated labs. We have developed a novel microFISH device to perform metaphase FISH on a chip which overcomes many shortcomings of the current laboratory protocols. This work...

  12. Metaphase FISH on a Chip: Miniaturized Microfluidic Device for Fluorescence in situ Hybridization

    Directory of Open Access Journals (Sweden)

    Niels Tommerup

    2010-11-01

    Full Text Available Fluorescence in situ Hybridization (FISH is a major cytogenetic technique for clinical genetic diagnosis of both inherited and acquired chromosomal abnormalities. Although FISH techniques have evolved and are often used together with other cytogenetic methods like CGH, PRINS and PNA-FISH, the process continues to be a manual, labour intensive, expensive and time consuming technique, often taking over 3–5 days, even in dedicated labs. We have developed a novel microFISH device to perform metaphase FISH on a chip which overcomes many shortcomings of the current laboratory protocols. This work also introduces a novel splashing device for preparing metaphase spreads on a microscope glass slide, followed by a rapid adhesive tape-based bonding protocol leading to rapid fabrication of the microFISH device. The microFISH device allows for an optimized metaphase FISH protocol on a chip with over a 20-fold reduction in the reagent volume. This is the first demonstration of metaphase FISH on a microfluidic device and offers a possibility of automation and significant cost reduction of many routine diagnostic tests of genetic anomalies.

  13. How eggs arrest at metaphase II: MPF stabilisation plus APC/C inhibition equals Cytostatic Factor

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    Jones Keith T

    2007-01-01

    Full Text Available Abstract Oocytes from higher chordates, including man and nearly all mammals, arrest at metaphase of the second meiotic division before fertilization. This arrest is due to an activity that has been termed 'Cytostatic Factor'. Cytostatic Factor maintains arrest through preventing loss in Maturation-Promoting Factor (MPF; CDK1/cyclin B. Physiologically, Cytostatic Factor – induced metaphase arrest is only broken by a Ca2+ rise initiated by the fertilizing sperm and results in degradation of cyclin B, the regulatory subunit of MPF through the Anaphase-Promoting Complex/Cyclosome (APC/C. Arrest at metaphase II may therefore be viewed as being maintained by inhibition of the APC/C, and Cytostatic Factor as being one or more pathways, one of which inhibits the APC/C, consorting in the preservation of MPF activity. Many studies over several years have implicated the c-Mos/MEK/MAPK pathway in the metaphase arrest of the two most widely studied vertebrates, frog and mouse. Murine downstream components of this cascade are not known but in frog involve members of the spindle assembly checkpoint, which act to inhibit the APC/C. Interesting these downstream components appear not to be involved in the arrest of mouse eggs, suggesting a lack of conservation with respect to c-Mos targets. However, the recent discovery of Emi2 as an egg specific APC/C inhibitor whose degradation is Ca2+ dependent has greatly increased our understanding of MetII arrest. Emi2 is involved in both the establishment and maintenance of metaphase II arrest in frog and mouse suggesting a conservation of metaphase II arrest. Its identity as the physiologically relevant APC/C inhibitor involved in Cytostatic Factor arrest prompted us to re-evaluate the role of the c-Mos pathway in metaphase II arrest. This review presents a model of Cytostatic Factor arrest, which is primarily induced by Emi2 mediated APC/C inhibition but which also requires the c-Mos pathway to set MPF levels within

  14. Metaphase chromosome analysis by ligation-mediated PCR: heritable chromatin structure and a comparison of active and inactive X chromosomes.

    Science.gov (United States)

    Hershkovitz, M; Riggs, A D

    1995-03-14

    We report that ligation-mediated PCR (LMPCR) can be used for high-resolution study of metaphase chromosomes, and we discuss the role of metaphase chromatin structure in the preservation of differentiated cell states. The X chromosome-linked human PGK1 (phosphoglycerate kinase 1) promoter region was investigated, and euchromatic active X chromosome (Xa) metaphase chromatin was compared with interphase Xa chromatin and to heterochromatic inactive X chromosome (Xi) metaphase and interphase chromatin. We find that (i) good-quality data at single-nucleotide resolution can be obtained by LMPCR analysis of dimethyl sulfate-treated intact metaphase cells; (ii) transcription factors present on the Xa promoter of interphase chromatin are not present on metaphase chromatin, establishing that the transcription complex on the PGK1 promoter must form de novo each cell generation; and (iii) the dimethyl sulfate reactivity pattern of Xa and Xi chromatin at metaphase is very similar to that of naked DNA. These results are discussed in the context of models for heritable chromatin structure and epigenetic mechanisms for cell memory, and they are also relevant to more general aspects of chromatin structure and differences between euchromatin and heterochromatin.

  15. Chromosome mapping by FISH to metaphase and interphase nuclei. Final report

    Energy Technology Data Exchange (ETDEWEB)

    Trask, B.

    1997-08-01

    The overall specific aims of this project were: (1) to determine the large-scale structure of interphase and metaphase chromosomes, in order to establish new capabilities for genome mapping by fluorescence in situ hybridization (FISH); (2) to detect chromosome abnormalities associated with genetic disease and map DNA sequences relative to them in order to facilitate the identification of new genes with disease-causing mutations; (3) to establish medium resolution physical maps of selected chromosomal regions using a combined metaphase and interphase mapping strategy and to corroborate physical and genetic maps and integrate these maps with the cytogenetic map; (4) to analyze the polymorphism and sequence evolution of subtelomeric regions of human chromosomes; (5) to establish a state-of-the-art FISH and image processing facility in the Department of Molecular Biotechnology, University of Washington, in order to map DNA sequences rapidly and accurately to benefit the Human Genome Project.

  16. Geant4.10 simulation of geometric model for metaphase chromosome

    Energy Technology Data Exchange (ETDEWEB)

    Rafat-Motavalli, L., E-mail: rafat@um.ac.ir; Miri-Hakimabad, H.; Bakhtiyari, E.

    2016-04-01

    In this paper, a geometric model of metaphase chromosome is explained. The model is constructed according to the packing ratio and dimension of the structure from nucleosome up to chromosome. A B-DNA base pair is used to construct 200 base pairs of nucleosomes. Each chromatin fiber loop, which is the unit of repeat, has 49,200 bp. This geometry is entered in Geant4.10 Monte Carlo simulation toolkit and can be extended to the whole metaphase chromosomes and any application in which a DNA geometrical model is needed. The chromosome base pairs, chromosome length, and relative length of chromosomes are calculated. The calculated relative length is compared to the relative length of human chromosomes.

  17. Ligation-mediated PCR for chromatin-structure analysis of interphase and metaphase chromatin.

    Science.gov (United States)

    Hershkovitz, M; Riggs, A D

    1997-02-01

    Chromatin structure is becoming increasingly recognized as important for a full understanding of gene function and cell memory. With regard to cell memory, which involves the transfer of chromatin-encoded epigenetic information from one cell generation to another, the detailed structure of metaphase chromatin is of crucial importance. In this paper we describe methods for the use of dimethyl sulfate, DNase I, and potassium permanganate for in vivo footprinting and chromatin analysis, with special emphasis on studies of metaphase cells. We review the use of ligation-mediated PCR for the analysis of chromatin, including the human phosphoglycerate kinase promoter, and also report initial studies of a matrix attachment region near the human beta-interferon gene.

  18. A potential tension-sensing mechanism that ensures timely anaphase onset upon metaphase spindle orientation.

    Science.gov (United States)

    Rajagopalan, Srividya; Bimbo, Andrea; Balasubramanian, Mohan K; Oliferenko, Snezhana

    2004-01-06

    The spindle orientation checkpoint (SOC) in fission yeast has been proposed to delay metaphase-to-anaphase transition when the spindle poles are misaligned with respect to the long axis of the cell. This checkpoint is activated in the absence of either an actomyosin division ring or astral microtubules. Although the SOC could be overridden in the absence of the transcription factor Atf1p, its mechanistic nature remained unclear. Here, we show that the SOC-triggered metaphase delay depends on a subset of the spindle assembly checkpoint (SAC) components Mph1p and Bub1p. Based on this finding and a detailed imaging of the spindle orientation process, we hypothesized that the spindle pole might contain proteins capable of sensing the achievement of spindle alignment. We identified the kendrin-like spindle pole body resident Pcp1p as a candidate molecule. A targeted mutation in its central domain specifically triggered the SOC in spite of the presence of oriented spindles, causing a metaphase delay that could be relieved in the absence of Mph1p, Bub1p, and Atf1p. Thus, Pcp1p might provide a link between the mechanical process of spindle alignment and the signal transduction that initiates anaphase.

  19. Stage-specific damage to synaptonemal complexes and metaphase chromosomes induced by X rays in male mouse germ cells

    International Nuclear Information System (INIS)

    Backer, L.C.; Sontag, M.R.; Allen, J.W.

    1991-01-01

    Synaptonemal complexes (SCs) reveal mutagen-induced effects in germ cell meiotic chromosomes. The study was aimed at characterizing relationships between SC and metaphase I chromosome damage following radiation exposure at various stages of spermatogenesis. Male mice were irradiated with doses of 0, 2, or 4 Gy, and spermatocytes were harvested at times consistent with earlier exposures as spermatogonial stem cells, preleptotene cells (premeiotic DNA synthesis), or meiotic prophase cells. After stem-cell exposure, twice as many rearrangements were observed in SCs as in metaphase I chromosomes. Irradiation during premeiotic DNA synthesis resulted in dose-related increases in SC breakage and rearrangements (including novel forms) and in metaphase chromosomal aberrations. Following prophase exposure, various types and levels of SC and metaphase damage were observed. Irradiation of zygotene cells led to high frequencies of chromosome multivalents in metaphase I without a correspondingly high level of damage in preceding prophase SCs. Thus, irradiation of premeiotic and meiotic cells results in variable relationships between SC and metaphase chromosome damage

  20. Metaphase I orientation of Robertsonian trivalents in the water-hyacinth grasshopper, Cornops aquaticum (Acrididae, Orthoptera

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    Pablo César Colombo

    2009-01-01

    Full Text Available Trivalents resulting from polymorphic Robertsonian rearrangements must have a regular orientation in metaphase I if the polymorphisms are to be maintained. It has been argued that redistribution of proximal and interstitial chiasmata to more distal positions is necessary for a convergent orientation, the only one that produces viable gametes. Cornops aquaticum is a South-American grasshopper that lives and feeds on water-hyacinths, and has three polymorphic Robertsonian rearrangements in its southernmost distribution area in Central Argentina and Uruguay. The orientation of trivalents in metaphase I, the formation of abnormal spermatids and the frequency and position of chiasmata in the trivalents, was analysed in a polymorphic population of C. aquaticus. In this study we observed a correlation between the number of trivalents with the frequency of abnormal spermatids; additionally, the number of chiasmata, especially proximal and interstitial ones, was strongly correlated with the frequency of the linear orientation. Therefore we confirmed our previous assumption, based on other evidence, that the chiasmata redistribution in fusion carriers is essential to the maintenance of the polymorphisms.

  1. Chromosomes in a genome-wise order: evidence for metaphase architecture.

    Science.gov (United States)

    Weise, Anja; Bhatt, Samarth; Piaszinski, Katja; Kosyakova, Nadezda; Fan, Xiaobo; Altendorf-Hofmann, Annelore; Tanomtong, Alongklod; Chaveerach, Arunrat; de Cioffi, Marcelo Bello; de Oliveira, Edivaldo; Walther, Joachim-U; Liehr, Thomas; Chaudhuri, Jyoti P

    2016-01-01

    One fundamental finding of the last decade is that, besides the primary DNA sequence information there are several epigenetic "information-layers" like DNA-and histone modifications, chromatin packaging and, last but not least, the position of genes in the nucleus. We postulate that the functional genomic architecture is not restricted to the interphase of the cell cycle but can also be observed in the metaphase stage, when chromosomes are most condensed and microscopically visible. If so, it offers the unique opportunity to directly analyze the functional aspects of genomic architecture in different cells, species and diseases. Another aspect not directly accessible by molecular techniques is the genome merged from two different haploid parental genomes represented by the homologous chromosome sets. Our results show that there is not only a well-known and defined nuclear architecture in interphase but also in metaphase leading to a bilateral organization of the two haploid sets of chromosomes. Moreover, evidence is provided for the parental origin of the haploid grouping. From our findings we postulate an additional epigenetic information layer within the genome including the organization of homologous chromosomes and their parental origin which may now substantially change the landscape of genetics.

  2. Drifter technique: a new method to obtain metaphases in Hep-2 cell line cultures

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    Eleonidas Moura Lima

    2005-07-01

    Full Text Available The Hep-2 cell line is derived from laryngeal carcinoma cells and is often utilized as a model in carcinogenesis and mutagenesis tests. To evaluate the proliferative potential of this line, we developed a cytogenetic methodology (drifter technique to obtain metaphases from cells that loose cellular adhesion when they underwent mitosis in culture. By this procedure, 2000 cells were counted, resulting in a mitotic index (MI of 22.2%. Although this MI was not statistically different from the one obtained using either a classical cytogenetic method or a cell synchronization technique, the drifter technique has the advantage of not requiring the use of some reagents for the obtention of metaphases and also of diminishing the consumption of maintenance reagents for this cell line.A linhagem celular Hep-2 é formada por células de carcinoma da laringe e é muito utilizada em modelos de carcinogênese e mutagenêse. Para avaliar o potencial proliferativo desta linhagem, desenvolvemos uma metodologia citogenética (técnica do sobrenadante para obtenção de metáfases a partir de células que, ao entrarem em mitose, perdem adesão celular, ficando em suspensão no meio de cultura. Através deste procedimento, foram contadas 2000 células, correspondendo a um índice mitótico (IM de 22.2% . Apesar de o IM obtido por esta técnica não ter sido estatisticamente diferente do IM obtido por outras metodologias citogenéticas clássicas, a técnica do sobrenadante é vantajosa porque elimina o uso de alguns reagentes utilizados na obtenção de metáfases e também diminui o consumo de reagentes de manutenção desta linhagem.

  3. Separate Location of Parental Chromosomes in Squashed Metaphases of Hybrid between Hordeum vulgare L. and Four Polyploid, Alien Species

    DEFF Research Database (Denmark)

    Jensen, J.; Linde-Laursen, Ib

    1984-01-01

    In 38 squashed, somatic metaphases of four hybrids between diploid Hordeum vulgare and two tetra-and two hexaploid alien species, each of the H. vulgare chromosomes was identifed, and differentiated from the chromosomes of the other parental species, by its Giemsa C-banding pattern. The H. vulgare...

  4. Detection of in-situ hybridization to human metaphase chromosomes by atomic force microscopy

    Science.gov (United States)

    Okamoto, Naoaki; Ishikawa, Mitsuru

    2000-04-01

    Detection of in situ hybridization to human metaphase chromosomes provides important information about gene mappings and about analysis of chromosomal disorders. We applied atomic force microscopy (AFM) to the detection of in situ hybridization to get better resolution as compared to light microscopy. Chromosomes were spread over a glass substrate and hybridized with DNA probes labeled with biotin or digoxigenin. The hybridized probes were reacted with streptavidin or anti-digoxigenin antibody, both of which were conjugated with 5-nm gold colloidal particles. We missed direct detection of the conjugated gold colloidal particles by micro-meter scale AFM scanning , but obtained clear topographic difference between the site of hybridization and the chromosome arm with the help of silver enhancement. We thus clearly detected the in situ hybridization using chromosome painting probes, alpha satellite probes, and locus specific gene probes by AFM. The in situ hybridization to DNA fiber was also detected by AFM. The detection of in situ hybridization by AFM has advantages over fluorescence in situ hybridization: no reduction of signal intensity under light irradiation. Application of AFM to the detection of in situ hybridization will be a useful method to analyze chromosomes.

  5. Single-nucleotide polymorphism array genotyping is equivalent to metaphase cytogenetics for diagnosis of Turner syndrome.

    Science.gov (United States)

    Prakash, Siddharth; Guo, Dongchuan; Maslen, Cheryl L; Silberbach, Michael; Milewicz, Dianna; Bondy, Carolyn A

    2014-01-01

    Turner syndrome is a developmental disorder caused by partial or complete monosomy for the X chromosome in 1 in 2,500 females. We hypothesized that single-nucleotide polymorphism (SNP) array genotyping could provide superior resolution in comparison to metaphase karyotype analysis to facilitate genotype-phenotype correlations. We genotyped 187 Turner syndrome patients with 733,000 SNP marker arrays. All cases met diagnostic criteria for Turner syndrome based on karyotypes (60%) or characteristic physical features. The SNP array results confirmed the diagnosis of Turner syndrome in 100% of cases. We identified a single X chromosome (45,X) in 113 cases. In 58 additional cases (31%), other mosaic cell lines were present, including isochromosomes (16%), rings (5%), and Xp deletions (8%). The remaining cases were mosaic for monosomy X and normal male or female cell lines. Array-based models of X chromosome structure were compatible with karyotypes in 104 of 116 comparable cases (90%). We found that the SNP array data did not detect X-autosome translocations (three cases) but did identify two derivative Y chromosomes and 13 large copy-number variants that were not detected by karyotyping. Our study is the first systematic comparison between the two methods and supports the utility of SNP array genotyping to address clinical and research questions in Turner syndrome.

  6. ERK3 is required for metaphase-anaphase transition in mouse oocyte meiosis.

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    Sen Li

    2010-09-01

    Full Text Available ERK3 (extracellular signal-regulated kinase 3 is an atypical member of the mitogen-activated protein (MAP kinase family of serine/threonine kinases. Little is known about its function in mitosis, and even less about its roles in mammalian oocyte meiosis. In the present study, we examined the localization, expression and functions of ERK3 during mouse oocyte meiotic maturation. Immunofluorescent analysis showed that ERK3 localized to the spindles from the pre-MI stage to the MII stage. ERK3 co-localized with α-tubulin on the spindle fibers and asters in oocytes after taxol treatment. Deletion of ERK3 by microinjection of ERK3 morpholino (ERK3 MO resulted in oocyte arrest at the MI stage with severely impaired spindles and misaligned chromosomes. Most importantly, the spindle assembly checkpoint protein BubR1 could be detected on kinetochores even in oocytes cultured for 10 h. Low temperature treatment experiments indicated that ERK3 deletion disrupted kinetochore-microtubule (K-MT attachments. Chromosome spreading experiments showed that knock-down of ERK3 prevented the segregation of homologous chromosomes. Our data suggest that ERK3 is crucial for spindle stability and required for the metaphase-anaphase transition in mouse oocyte maturation.

  7. Dynein Light Intermediate Chain 2 Facilitates the Metaphase to Anaphase Transition by Inactivating the Spindle Assembly Checkpoint.

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    Sagar P Mahale

    Full Text Available The multi-functional molecular motor cytoplasmic dynein performs diverse essential roles during mitosis. The mechanistic importance of the dynein Light Intermediate Chain homologs, LIC1 and LIC2 is unappreciated, especially in the context of mitosis. LIC1 and LIC2 are believed to exist in distinct cytoplasmic dynein complexes as obligate subunits. LIC1 had earlier been reported to be required for metaphase to anaphase progression by inactivating the kinetochore-microtubule attachment-sensing arm of the spindle assembly checkpoint (SAC. However, the functional importance of LIC2 during mitosis remains elusive. Here we report prominent novel roles for the LIC2 subunit of cytoplasmic dynein in regulating the spindle assembly checkpoint. LIC2 depletion in mammalian cells led to prolonged metaphase arrest in the presence of an active SAC and also to stretched kinetochores, thus implicating it in SAC inactivation. Quantitative fluorescence microscopy of SAC components revealed accumulation of both attachment- and tension-sensing checkpoint proteins at metaphase kinetochores upon LIC2 depletion. These observations support a stronger and more diverse role in checkpoint inactivation for LIC2 in comparison to its close homolog LIC1. Our study uncovers a novel functional hierarchy during mitotic checkpoint inactivation between the closely related but homologous LIC subunits of cytoplasmic dynein. These subtle functional distinctions between dynein subpopulations could be exploited to study specific aspects of the spindle assembly checkpoint, which is a key mediator of fidelity in eukaryotic cell division.

  8. Analysis of the phospholipid profile of metaphase II mouse oocytes undergoing vitrification.

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    Jaehun Jung

    Full Text Available Oocyte freezing confers thermal and chemical stress upon the oolemma and various other intracellular structures due to the formation of ice crystals. The lipid profiles of oocytes and embryos are closely associated with both, the degrees of their membrane fluidity, as well as the degree of chilling and freezing injuries that may occur during cryopreservation. In spite of the importance of lipids in the process of cryopreservation, the phospholipid status in oocytes and embryos before and after freezing has not been investigated. In this study, we employed mass spectrometric analysis to examine if vitrification has an effect on the phospholipid profiles of mouse oocytes. Freshly prepared metaphase II mouse oocytes were vitrified using copper grids and stored in liquid nitrogen for 2 weeks. Fresh and vitrified-warmed oocytes were subjected to phospholipid extraction procedure. Mass spectrometric analyses revealed that multiple species of phospholipids are reduced in vitrified-warmed oocytes. LIFT analyses identified 31 underexpressed and 5 overexpressed phospholipids in vitrified mouse oocytes. The intensities of phosphatidylinositol (PI {18∶2/16∶0} [M-H]- and phosphatidylglycerol (PG {14∶0/18∶2} [M-H]- were decreased the most with fold changes of 30.5 and 19.1 in negative ion mode, respectively. Several sphingomyelins (SM including SM {d38∶3} [M+H]+ and SM {d34∶0} [M+K]+ were decreased significantly in positive ion mode. Overall, the declining trend of multiple phospholipids demonstrates that vitrification has a marked effect on phospholipid profiles of oocytes. These results show that the identified phospholipids can be used as potential biomarkers of oocyte undergoing vitrification and will allow for the development of strategies to preserve phospholipids during oocyte cryopreservation.

  9. Cyclophosphamide and acrolein induced oxidative stress leading to deterioration of metaphase II mouse oocyte quality.

    Science.gov (United States)

    Jeelani, Roohi; Khan, Sana N; Shaeib, Faten; Kohan-Ghadr, Hamid-Reza; Aldhaheri, Sarah R; Najafi, Tohid; Thakur, Mili; Morris, Robert; Abu-Soud, Husam M

    2017-09-01

    Cyclophosphamide (CTX) is a chemotherapeutic agent widely used to treat ovarian, breast, and hematological cancers as well as autoimmune disorders. Such chemotherapy is associated with reproductive failure and premature ovarian insufficiency. The mechanism by which CTX and/or its main metabolite, acrolein, affect female fertility remains unclear, but it is thought to be caused by an overproduction of reactive oxygen species (ROS). Here, we investigated the effect of CTX on metaphase II mouse oocytes obtained from treated animals (120mg/kg, 24h of single treatment), and oocytes directly exposed to increasing concentrations of CTX and acrolein (n=480; 0, 5, 10, 25, 50, and 100μM) with and without cumulus cells (CCs) for 45min which correlates to the time of maximum peak plasma concentrations after administration. Oocytes were fixed and subjected to indirect immunofluorescence and were scored based on microtubule spindle structure (MT) and chromosomal alignment (CH). Generation of ROS was evaluated using the Cellular Reactive Oxygen Species Detection Assay Kit. Deterioration of oocyte quality was noted when oocytes were obtained from CTX treated mice along with CTX and acrolein treated oocytes in a dose-dependent manner as shown by an increase in poor scores. Acrolein had an impact at a significantly lower level as compared to CTX, plateau at 10μM versus 50μM, respectively. These variation is are associated with the higher amount of ROS generated with acrolein exposure as compared to CTX (pacrolein scavengers may mitigate the damaging effects of these compounds and help women undergoing such treatment. Copyright © 2017. Published by Elsevier Inc.

  10. Detection of short repeated genomic sequences on metaphase chromosomes using padlock probes and target primed rolling circle DNA synthesis

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    Stougaard Magnus

    2007-11-01

    Full Text Available Abstract Background In situ detection of short sequence elements in genomic DNA requires short probes with high molecular resolution and powerful specific signal amplification. Padlock probes can differentiate single base variations. Ligated padlock probes can be amplified in situ by rolling circle DNA synthesis and detected by fluorescence microscopy, thus enhancing PRINS type reactions, where localized DNA synthesis reports on the position of hybridization targets, to potentially reveal the binding of single oligonucleotide-size probe molecules. Such a system has been presented for the detection of mitochondrial DNA in fixed cells, whereas attempts to apply rolling circle detection to metaphase chromosomes have previously failed, according to the literature. Methods Synchronized cultured cells were fixed with methanol/acetic acid to prepare chromosome spreads in teflon-coated diagnostic well-slides. Apart from the slide format and the chromosome spreading everything was done essentially according to standard protocols. Hybridization targets were detected in situ with padlock probes, which were ligated and amplified using target primed rolling circle DNA synthesis, and detected by fluorescence labeling. Results An optimized protocol for the spreading of condensed metaphase chromosomes in teflon-coated diagnostic well-slides was developed. Applying this protocol we generated specimens for target primed rolling circle DNA synthesis of padlock probes recognizing a 40 nucleotide sequence in the male specific repetitive satellite I sequence (DYZ1 on the Y-chromosome and a 32 nucleotide sequence in the repetitive kringle IV domain in the apolipoprotein(a gene positioned on the long arm of chromosome 6. These targets were detected with good efficiency, but the efficiency on other target sites was unsatisfactory. Conclusion Our aim was to test the applicability of the method used on mitochondrial DNA to the analysis of nuclear genomes, in particular as

  11. The Aurora Kinase in Trypanosoma brucei plays distinctive roles in metaphase-anaphase transition and cytokinetic initiation.

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    Ziyin Li

    2009-09-01

    Full Text Available Aurora B kinase is an essential regulator of chromosome segregation with the action well characterized in eukaryotes. It is also implicated in cytokinesis, but the detailed mechanism remains less clear, partly due to the difficulty in separating the latter from the former function in a growing cell. A chemical genetic approach with an inhibitor of the enzyme added to a synchronized cell population at different stages of the cell cycle would probably solve this problem. In the deeply branched parasitic protozoan Trypanosoma brucei, an Aurora B homolog, TbAUK1, was found to control both chromosome segregation and cytokinetic initiation by evidence from RNAi and dominant negative mutation. To clearly separate these two functions, VX-680, an inhibitor of TbAUK1, was added to a synchronized T. brucei procyclic cell population at different cell cycle stages. The unique trans-localization pattern of the chromosomal passenger complex (CPC, consisting of TbAUK1 and two novel proteins TbCPC1 and TbCPC2, was monitored during mitosis and cytokinesis by following the migration of the proteins tagged with enhanced yellow fluorescence protein in live cells with time-lapse video microscopy. Inhibition of TbAUK1 function in S-phase, prophase or metaphase invariably arrests the cells in the metaphase, suggesting an action of TbAUK1 in promoting metaphase-anaphase transition. TbAUK1 inhibition in anaphase does not affect mitotic exit, but prevents trans-localization of the CPC from the spindle midzone to the anterior tip of the new flagellum attachment zone for cytokinetic initiation. The CPC in the midzone is dispersed back to the two segregated nuclei, while cytokinesis is inhibited. In and beyond telophase, TbAUK1 inhibition has no effect on the progression of cytokinesis or the subsequent G1, S and G2 phases until a new metaphase is attained. There are thus two clearly distinct points of TbAUK1 action in T. brucei: the metaphase-anaphase transition and

  12. The dormant and the fully competent oocyte: comparing the transcriptome of human oocytes from primordial follicles and in metaphase II

    DEFF Research Database (Denmark)

    Grøndahl, Marie Louise; Borup, Rehannah; Vikeså, Jonas

    2013-01-01

    Oocytes become enclosed in primordial follicles during fetal life and remain dormant there until activation followed by growth and meiotic resumption. Current knowledge about the molecular pathways involved in oogenesis is incomplete. This study identifies the specific transcriptome of the human...... oocyte in the quiescent state and at the pinnacle of maturity at ovulation. In silico bioinformatic comparisons were made between the transcriptome of human oocytes from dormant primordial follicles and that of human metaphase II (MII) oocytes and granulosa cells and unique gene expression profiles were...

  13. ADP-ribosylation of nonhistone proteins from metaphase and interphase HeLa cells: factors responsible for differences

    International Nuclear Information System (INIS)

    Adolph, K.W.

    1986-01-01

    A striking reduction was previously detected for HeLa metaphase chromosomes, compared to interphase nuclei, in the number of modified nonhistone species. Several factors which could contribute to this cell cycle change in ADP-ribosylation have therefore been examined. In these experiments, mitotic or interphase cells were incubated with [ 32 P]NAD, chromosomes and nuclei were prepared, and the proteins were resolved by polyacrylamide gel electrophoresis. The level of incorporation of 32 P label was found to be substantially influenced by chromosome expansion, DNA nicking, disruption of chromosomes or nuclei, and the growth activity of cells. The level of ADP-ribosylation was not greatly affected by the presence of inhibitors of RNA, DNA, and protein synthesis. NAD concentration influenced the extent of labelling but not the pattern of labeled species. A similar change in the pattern from interphase to mitosis was observed for whole cells as well as for isolated chromosomes and nuclei. The procedure used to arrest cells in mitosis was not artifactually responsible for the results. The difference in metaphase and interphase ADP-ribosylation is not confined to HeLa cells, since comparable patterns were found for chromosomes and nuclei from Novikoff rat hepatoma cells

  14. Thiol protease-specific inhibitor E-64 arrests human epidermoid carcinoma A431 cells at mitotic metaphase

    International Nuclear Information System (INIS)

    Shoji-Kasai, Y.; Senshu, M.; Iwashita, S.; Imahori, K.

    1988-01-01

    E-64-d /ethyl (2S, 3S)-3-[(S)-3-methyl-1-(3-methylbutylcarbamoyl)butylcarbamoyl]oxirane-2-carboxylate/, a membrane-permeant derivative of the thiol protease-specific inhibitor E-64, was found to arrest human epidermoid carcinoma A431 cells at mitotic metaphase. This effect was dose-dependent with a threshold of 20μg/ml in chemically defined culture medium. Cell cycle analysis by flow cytometry showed that the relative proportion of the G 2 /M population increased 2.5-fold after treatment of the cells with E-64 (100 μg/ml) for 5 hr. In addition, time-lapse video analysis showed that E-64-treated cells remained at metaphase for an extended period after rounding-up, whereas untreated cells were able to complete mitosis within 42.0 +/- 5.7 min. Some treated cells were able to complete mitosis, while others did not do so within limits of the authors observation. An approach to the molecular basis of this phenomenon, they have shown that several cellular proteins can be labeled by incubation of cells with radioactive E-64-d

  15. Automated identification of abnormal metaphase chromosome cells for the detection of chronic myeloid leukemia using microscopic images

    Science.gov (United States)

    Wang, Xingwei; Zheng, Bin; Li, Shibo; Mulvihill, John J.; Chen, Xiaodong; Liu, Hong

    2010-07-01

    Karyotyping is an important process to classify chromosomes into standard classes and the results are routinely used by the clinicians to diagnose cancers and genetic diseases. However, visual karyotyping using microscopic images is time-consuming and tedious, which reduces the diagnostic efficiency and accuracy. Although many efforts have been made to develop computerized schemes for automated karyotyping, no schemes can get be performed without substantial human intervention. Instead of developing a method to classify all chromosome classes, we develop an automatic scheme to detect abnormal metaphase cells by identifying a specific class of chromosomes (class 22) and prescreen for suspicious chronic myeloid leukemia (CML). The scheme includes three steps: (1) iteratively segment randomly distributed individual chromosomes, (2) process segmented chromosomes and compute image features to identify the candidates, and (3) apply an adaptive matching template to identify chromosomes of class 22. An image data set of 451 metaphase cells extracted from bone marrow specimens of 30 positive and 30 negative cases for CML is selected to test the scheme's performance. The overall case-based classification accuracy is 93.3% (100% sensitivity and 86.7% specificity). The results demonstrate the feasibility of applying an automated scheme to detect or prescreen the suspicious cancer cases.

  16. Direct fluorescence in situ hybridization on human metaphase chromosomes using quantum dot-platinum labeled DNA probes

    Energy Technology Data Exchange (ETDEWEB)

    Hwang, Gyoyeon [Chemical Kinomics Research Center, Future Convergence Research Division, Korea Institute of Science and Technology, Hwarangno 14-gil 5, Seongbuk-gu, Seoul 136-791 (Korea, Republic of); Biological Chemistry, Korea University of Science and Technology, 217, Gajeong-ro, Yuseong-gu, Deajeon (Korea, Republic of); Lee, Hansol [Chemical Kinomics Research Center, Future Convergence Research Division, Korea Institute of Science and Technology, Hwarangno 14-gil 5, Seongbuk-gu, Seoul 136-791 (Korea, Republic of); Lee, Jiyeon, E-mail: jylee@kist.re.kr [Chemical Kinomics Research Center, Future Convergence Research Division, Korea Institute of Science and Technology, Hwarangno 14-gil 5, Seongbuk-gu, Seoul 136-791 (Korea, Republic of); Biological Chemistry, Korea University of Science and Technology, 217, Gajeong-ro, Yuseong-gu, Deajeon (Korea, Republic of)

    2015-11-13

    The telomere shortening in chromosomes implies the senescence, apoptosis, or oncogenic transformation of cells. Since detecting telomeres in aging and diseases like cancer, is important, the direct detection of telomeres has been a very useful biomarker. We propose a telomere detection method using a newly synthesized quantum dot (QD) based probe with oligonucleotide conjugation and direct fluorescence in situ hybridization (FISH). QD-oligonucleotides were prepared with metal coordination bonding based on platinum-guanine binding reported in our previous work. The QD-oligonucleotide conjugation method has an advantage where any sequence containing guanine at the end can be easily bound to the starting QD-Pt conjugate. A synthesized telomeric oligonucleotide was bound to the QD-Pt conjugate successfully and this probe hybridized specifically on the telomere of fabricated MV-4-11 and MOLT-4 chromosomes. Additionally, the QD-telomeric oligonucleotide probe successfully detected the telomeres on the CGH metaphase slide. Due to the excellent photostability and high quantum yield of QDs, the QD-oligonucleotide probe has high fluorescence intensity when compared to the organic dye-oligonucleotide probe. Our QD-oligonucleotide probe, conjugation method of this QD probe, and hybridization protocol with the chromosomes can be a useful tool for chromosome painting and FISH. - Highlights: • We prepared a probe linked between QD and telomeric oligonucleotide with platinum-guanine bonding. • Telomeres were detected by our new telomere probes successfully in three different human metaphase chromosomes. • QDPt-DNA probe has high fluorescence intensity in comparison with organic dye-DNA probe.

  17. Correlation between DNA ploidy, metaphase high-resolution comparative genomic hybridization results and clinical outcome of synovial sarcoma

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    Papp Gergő

    2011-11-01

    Full Text Available Abstract Background Although synovial sarcoma is the 3rd most commonly occurring mesenchymal tumor in young adults, usually with a highly aggressive clinical course; remarkable differences can be seen regarding the clinical outcome. According to comparative genomic hybridization (CGH data published in the literature, the simple and complex karyotypes show a correlation between the prognosis and clinical outcome. In addition, the connection between DNA ploidy and clinical course is controversial. The aim of this study was using a fine-tuning interpretation of our DNA ploidy results and to compare these with metaphase high-resolution CGH (HR-CGH results. Methods DNA ploidy was determined on Feulgen-stained smears in 56 synovial sarcoma cases by image cytometry; follow up was available in 46 cases (average: 78 months. In 9 cases HR-CGH analysis was also available. Results 10 cases were found DNA-aneuploid, 46 were DNA-diploid by image cytometry. With fine-tuning of the diploid cases according to the 5c exceeding events (single cell aneuploidy, 33 cases were so called "simple-diploid" (without 5c exceeding events and 13 cases were "complex-diploid"; containing 5c exceeding events (any number. Aneuploid tumors contained large numbers of genetic alterations with the sum gain of at least 2 chromosomes (A-, B- or C-group detected by HR-CGH. In the "simple-diploid" cases no or few genetic alterations could be detected, whereas the "complex-diploid" samples numerous aberrations (equal or more than 3 could be found. Conclusions Our results show a correlation between the DNA-ploidy, a fine-tuned DNA-ploidy and the HR-CGH results. Furthermore, we found significant correlation between the different ploidy groups and the clinical outcome (p

  18. Securin associates with APCCdh1 in prometaphase but its destruction is delayed by Rae1 and Nup98 until the metaphase/anaphase transition.

    Science.gov (United States)

    Jeganathan, Karthik B; Baker, Darren J; van Deursen, Jan M

    2006-02-01

    Precisely timed ubiquitin-mediated proteolysis of mitotic regulators by the anaphase-promoting complex (APC) governs the orderly passage of cells through mitosis. The established view is that Cdc20-activated APC (APC(Cdc20)) mediates the destruction of cyclin B and securin at the metaphase/anaphase transition, and that Cdh1-activated APC (APC(Cdh1)) has no role in this process. We recently reported that securin, but not cyclin B, is prematurely targeted for destruction by the APC in mutant mice that have low levels of the nuclear transport factors Rae1 and Nup98. We found that Rae1 and Nup98 assemble into a complex with APC(Cdh1) in prometaphase and act to delay APC(Cdh1)-mediated ubiquitination of securin until the metaphase/anaphase transition. Here we show that Rae1 and Nup98 not only form a complex with APC(Cdh1) in prometaphase but also with securin. This finding suggests that the Rae1-Nup98 complex does not inhibit early destruction of securin by preventing APC(Cdh1) from binding to securin, but by preventing ubiquitination of APC(Cdh1)-bound securin. We propose that the formation of APC(Cdh1)-securin complexes in prometaphase primes the cell for rapid securin degradation after release of the inhibitory Rae1-Nup98 complex at the metaphase/anaphase transition. We further report here that mutant mice with low levels of the Rae1-Nup98 complex are not prone to develop spontaneous tumors, despite massive aneuploidy. However, Rae1/Nup98 mutant mice are significantly more susceptible to DMBA-induced lung tumors than wild-type mice, indicating that combined Rae1/ Nup98 haplo-insufficiency does promote tumorigenesis when certain cancer-critical genes are also mutated.

  19. Single-Cell Dynamic Analysis of Mitosis in Haploid Embryonic Stem Cells Shows the Prolonged Metaphase and Its Association with Self-diploidization.

    Science.gov (United States)

    Guo, Ao; Huang, Shichao; Yu, Jiali; Wang, Huihan; Li, Haisen; Pei, Gang; Shen, Li

    2017-05-09

    The recent establishment of mammalian haploid embryonic stem cells (ESCs) provides new possibilities for genetic screening and for understanding genome evolution and function. However, the dynamics of mitosis in haploid ESCs is still unclear. Here, we report that the duration of mitosis in haploid ESCs, especially the metaphase, is significantly longer than that in diploid ESCs. Delaying mitosis by chemicals increased self-diploidization of haploid ESCs, while shortening mitosis stabilized haploid ESCs. Taken together, our study suggests that the delayed mitosis of haploid ESCs is associated with self-diploidization. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  20. Single-Cell Dynamic Analysis of Mitosis in Haploid Embryonic Stem Cells Shows the Prolonged Metaphase and Its Association with Self-diploidization

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    Ao Guo

    2017-05-01

    Full Text Available The recent establishment of mammalian haploid embryonic stem cells (ESCs provides new possibilities for genetic screening and for understanding genome evolution and function. However, the dynamics of mitosis in haploid ESCs is still unclear. Here, we report that the duration of mitosis in haploid ESCs, especially the metaphase, is significantly longer than that in diploid ESCs. Delaying mitosis by chemicals increased self-diploidization of haploid ESCs, while shortening mitosis stabilized haploid ESCs. Taken together, our study suggests that the delayed mitosis of haploid ESCs is associated with self-diploidization.

  1. A noncoding RNA containing a SINE-B1 motif associates with meiotic metaphase chromatin and has an indispensable function during spermatogenesis.

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    Ryusuke Nakajima

    Full Text Available A search for early response genes that are activated following germ cell induction from mouse embryonic stem cells in vitro led us to the isolation of a long noncoding RNA that contains a SINE (short interspersed element-B1F motif that was named R53. In situ hybridization and northern blot analyses revealed that the R53 subfragment RNA bears a B1F motif, is processed from the primary transcript, is expressed in adult testis and is predominantly localized in meiotic metaphase chromatin during spermatogenesis. Recent studies of chromosome-associated RNAs have explored novel functions of noncoding RNAs. Specifically, chromosome-bound noncoding RNAs function not only as structural components of chromosome but also as scaffolds that recruit epigenetic modulators for transcriptional regulation, and they are dynamically rearranged during the cell cycle. However, few studies have explored meiotic chromatin; thus, R53 RNA appears to be the first long noncoding RNA to be tightly associated with the metaphase chromatin during spermatogenesis. Furthermore, R53 knockdown using a lentivirus-mediated RNAi injected into mouse testis and organ culture of the fragments revealed a remarkable reduction in postmeiotic cells and irregular up-regulation of several postmeiotic genes, which suggests the possibility that the SINE-B1-derived noncoding RNA R53 plays an indispensable role in the transcriptional regulation of key spermatogenesis genes.

  2. ''Protective'' effect of cells gamma-irradiation at the metaphase of mitosis after UV-irradiation at the S-period

    International Nuclear Information System (INIS)

    Lebedeva, L.I.; Chubykin, V.L.

    1975-01-01

    As a result of the ultraviolet irradiation in vitro of the embryo fibroblasts of BALB mice in the S-stage with an incident dose of 40 erg/mm 2 , 20.1% cells showed chromosome aberrations. Additional gamma irradiation of cells in the metaphase of the first mitosis with a dose of 5 krad leads with a high degree of certainty to a decrease to 11.7% in the frequency of aberrant cells observed in the same mitotic stage. The frequency of spontaneous aberrations does not change during the first few minutes after the gamma irradiation of intact cells. The ''protective'' effect of gamma rays cannot be attributed to non-uniform changes in the duration of the mitotic stages for aberrant and normal cells, to the adhesion of chromosome fragments or to the breaking of bridges in the anaphase. The destruction of cells during irradiation is also an unlikely explanation of the observed effect. It is assumed that the decrease in the frequency of aberrations is a result of the previously predicted modification of the processes involved, when potential chromosome damage becomes visible abberations during metaphase. (auth.)

  3. [Tripartite motif-containing protein 34 (TRIM34) colocalized with micronuclei chromosome and hampers its movement to equatorial plate during the metaphase stage of mitosis].

    Science.gov (United States)

    Sun, Dakang; An, Xinye; Ji, Bing; Cheng, Yanli; Gao, Honglian; Tian, Mingming

    2016-06-01

    Objective To examine whether tripartite motif-containing protein 34 (TRIM34) is colocalized with micronuclei and investigate the influence on the movement of micronuclei chromosome in mitosis. Methods The eukaryotic expression vector TRIM34-pEGFP-N3 was constructed, identified and then transfected into HEK293T cells. With 4', 6-diamidino-2-phenylindole 2HCI (DAPI) staining, the colocalization between TRIM34 and micronuclei was observed under a fluorescence microscope. Moreover, MitoTracker(R)Deep Red was used to identify the colocalization between the complex of TRIM34-micronulei and mitochondria under a confocal microscope. Finally, the effect of TRIM34 on the movement of micronuclei chromosome in mitosis was examined. Results DNA sequencing confirmed that the vector TRIM34-pEGFP-N3 was constructed successfully. A fluorescence microscope revealed that TRIM34 could be colocalized with micronuclei in HEK293T cells transfected with TRIM34-pEGFP-N3. In the same manner, a confocal microscope distinctly showed that TRIM34 was colocalized with micronuclei similarly in appearance. However, there was no distinguished colocalization relationship between the complex of TRIM34-micronulei and mitochondria. Interestingly, the micronuclei chromosome conjugated with TRIM34 was hardly transferred to equatorial plate during the metaphase stage of mitosis. Conclusion TRIM34 is colocalized with micronuclei chromosome and hampers its movement to equatorial plate in mitosis.

  4. The G-protein regulatory (GPR) motif-containing Leu-Gly-Asn-enriched protein (LGN) and Gialpha3 influence cortical positioning of the mitotic spindle poles at metaphase in symmetrically dividing mammalian cells.

    Science.gov (United States)

    Blumer, Joe B; Kuriyama, Ryoko; Gettys, Thomas W; Lanier, Stephen M

    2006-12-01

    We addressed the role of the G-protein regulatory (GPR) motif-containing Leu-Gly-Asn-enriched protein (LGN) and G-proteins (Gialpha3) in the positioning of the spindle pole during mammalian cell division. Immunocytochemistry indicated that both LGN and Gialpha3 co-localized at the spindle pole and at the midbody and the cell cortex during the different phases of mitosis. In marked contrast to the positioning of the spindle pole at metaphase midway between the cell cortex and the metaphase plate, the spindle pole was juxtaposed with the cell cortex at metaphase following increased expression of Gialpha3 and LGN. This repositioning of the spindle pole required the interaction of LGN with Gialpha. The influence of LGN and Gialpha3 on the cortical positioning of the spindle pole likely reflects either stronger pulling forces on the spindle pole exerted from the cell cortex or increased pushing forces exerted on the spindle pole from the mitotic spindle indicating that these events are regulated by GPR motif-containing proteins and G-proteins independent of asymmetry.

  5. 45S rDNA regions are chromosome fragile sites expressed as gaps in vitro on metaphase chromosomes of root-tip meristematic cells in Lolium spp.

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    Jing Huang

    Full Text Available BACKGROUND: In humans, chromosome fragile sites are regions that are especially prone to forming non-staining gaps, constrictions or breaks in one or both of the chromatids on metaphase chromosomes either spontaneously or following partial inhibition of DNA synthesis and have been well identified. So far, no plant chromosome fragile sites similar to those in human chromosomes have been reported. METHODS AND RESULTS: During the course of cytological mapping of rDNA on ryegrass chromosomes, we found that the number of chromosomes plus chromosome fragments was often more than the expected 14 in most cells for Lolium perenne L. cv. Player by close cytological examination using a routine chromosome preparation procedure. Further fluorescent in situ hybridization (FISH using 45S rDNA as a probe indicated that the root-tip cells having more than a 14-chromosome plus chromosome fragment count were a result of chromosome breakage or gap formation in vitro (referred to as chromosome lesions at 45S rDNA sites, and 86% of the cells exhibited chromosome breaks or gaps and all occurred at the sites of 45S rDNA in Lolium perenne L. cv. Player, as well as in L. multiflorum Lam. cv. Top One. Chromatin depletion or decondensation occurred at various locations within the 45S rDNA regions, suggesting heterogeneity of lesions of 45S rDNA sites with respect to their position within the rDNA region. CONCLUSIONS: The chromosome lesions observed in this study are very similar cytologically to that of fragile sites observed in human chromosomes, and thus we conclude that the high frequency of chromosome lesions in vitro in Lolium species is the result of the expression of 45S rDNA fragile sites. Possible causes for the spontaneous expression of fragile sites and their potential biological significance are discussed.

  6. Mouse oocytes fertilised by ICSI during in vitro maturation retain the ability to be activated after refertilisation in metaphase II and can generate Ca2+ oscillations

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    Pomorski Paweł

    2007-06-01

    Full Text Available Abstract Background At fertilisation, mammalian oocytes are activated by oscillations of intracellular Ca2+ ([Ca2+]i. Phospholipase Cζ, which is introduced by fertilising spermatozoon, triggers [Ca2+]i oscillations through the generation of inositol 1,4,5-triphosphate (IP3, which causes Ca2+ release by binding to IP3 receptors located on the endoplasmic reticulum (ER of the oocyte. Ability to respond to this activating stimulus develops during meiotic maturation of the oocyte. Here we examine how the development of this ability is perturbed when a single spermatozoon is introduced into the oocyte prematurely, i.e. during oocyte maturation. Results Mouse oocytes during maturation in vitro were fertilised by ICSI (intracytoplasmic sperm injection 1 – 4 h after germinal vesicle break-down (GVBD and were subsequently cultured until they reached metaphase II (MII stage. At MII stage they were fertilised in vitro for the second time (refertilisation. We observed that refertilised oocytes underwent activation with similar frequency as control oocytes, which also went through maturation in vitro, but were fertilised only once at MII stage (87% and 93%, respectively. Refertilised MII oocytes were able to develop [Ca2+]i oscillations in response to penetration by spermatozoa. We found however, that they generated a lower number of transients than control oocytes. We also showed that the oocytes, which were fertilised during maturation had a similar level of MPF activity as control oocytes, which were not subjected to ICSI during maturation, but had reduced level of IP3 receptors. Conclusion Mouse oocytes, which were experimentally fertilised during maturation retain the ability to generate repetitive [Ca2+]i transients, and to be activated after completion of maturation.

  7. A comparative analysis of chromosome pairing at metaphase I in interspecific hybrids between durum wheat (Triticum turgidum L.) and the most widespread Aegilops species.

    Science.gov (United States)

    Cifuentes, M; Garcia-Agüero, V; Benavente, E

    2010-07-01

    Homoeologous metaphase I (MI) associations in hybrids between durum wheat and its wild allotetraploid relatives Aegilops neglecta, Ae. triuncialis and Ae. ventricosa have been characterized by a genomic in situ hybridization procedure that allows simultaneous discrimination of A, B and wild species genomes. Earlier results in equivalent hybrids with the wild species Ae. cylindrica and Ae. geniculata have also been considered to comparatively assay the MI pairing pattern of the durum wheat x Aegilops interspecific combinations more likely to occur in nature. The general picture can be drawn as follows. A and B wheat genomes pair with each other less than the 2 wild constituent genomes do in any of the hybrid combinations examined. Interspecific wheat-wild associations account for 60-70% of total MI pairing in all hybrids, except in that derived from Ae. triuncialis, but the A genome is always the wheat partner most frequently involved in MI pairing with the wild homoeologues. Hybrids with Ae. cylindrica, Ae. geniculata and Ae. ventricosa showed similar reduced levels of MI association and virtually identical MI pairing patterns. However, certain recurrent differences were found when the pattern of homoeologous pairing of hybrids from either Ae. triuncialis or Ae. neglecta was contrasted to that observed in the other durum wheat hybrid combinations. In the former case, a remarkable preferential pairing between the wild species constituent genomes U(t) and C(t) seems to be the reason, whereas a general promotion of homoeologous pairing, qualitatively similar to that observed under the effect of the ph1c mutation, appears to occur in the hybrid with Ae. neglecta. It is further discussed whether the results reported here can be extrapolated to the corresponding bread wheat hybrid combinations. Copyright 2010 S. Karger AG, Basel.

  8. Separase Cleaves the N-Tail of the CENP-A Related Protein CPAR-1 at the Meiosis I Metaphase-Anaphase Transition in C. elegans.

    Directory of Open Access Journals (Sweden)

    Joost Monen

    Full Text Available Centromeres are defined epigenetically in the majority of eukaryotes by the presence of chromatin containing the centromeric histone H3 variant CENP-A. Most species have a single gene encoding a centromeric histone variant whereas C. elegans has two: HCP-3 (also known as CeCENP-A and CPAR-1. Prior RNAi replacement experiments showed that HCP-3 is the functionally dominant isoform, consistent with CPAR-1 not being detectable in embryos. GFP::CPAR-1 is loaded onto meiotic chromosomes in diakinesis and is enriched on bivalents until meiosis I. Here we show that GFP::CPAR-1 signal loss from chromosomes precisely coincides with homolog segregation during anaphase I. This loss of GFP::CPAR-1 signal reflects proteolytic cleavage between GFP and the histone fold of CPAR-1, as CPAR-1::GFP, in which GFP is fused to the C-terminus of CPAR-1, does not exhibit any loss of GFP signal. A focused candidate screen implicated separase, the protease that initiates anaphase by cleaving the kleisin subunit of cohesin, in this cleavage reaction. Examination of the N-terminal tail sequence of CPAR-1 revealed a putative separase cleavage site and mutation of the signature residues in this site eliminated the cleavage reaction, as visualized by retention of GFP::CPAR-1 signal on separating homologous chromosomes at the metaphase-anaphase transition of meiosis I. Neither cleaved nor uncleavable CPAR-1 were centromere-localized in mitosis and instead localized throughout chromatin, indicating that centromere activity has not been retained in CPAR-1. Although the functions of CPAR-1 and of its separase-dependent cleavage remain to be elucidated, this effort reveals a new substrate of separase and provides an in vivo biosensor to monitor separase activity at the onset of meiosis I anaphase.

  9. DNA Catenation Maintains Structure of Human Metaphase Chromosomes

    DEFF Research Database (Denmark)

    L. V. Bauer, David; Marie, Rodolphe; Rasmussen, Kristian Hagsted

    2012-01-01

    -on-a-chip microfluidic device and fluorescence microscopy, coupled with a simple image analysis pipeline, to digest chromosomal proteins and examine the structure of the remaining DNA, which maintains the canonical ‘X’ shape. By directly staining DNA, we observe that DNA catenation between sister chromatids (separated...... by fluid flow) is composed of distinct fibres of DNA concentrated at the centromeres. Disrupting the catenation of the chromosomes with Topoisomerase IIa significantly alters overall chromosome shape, suggesting that DNA catenation must be simultaneously maintained for correct chromosome condensation...

  10. Metaphase chromosome aberrations as markers of radiation exposure and dose

    Energy Technology Data Exchange (ETDEWEB)

    Brooks, A.L.; Khan, M.A.; Jostes, R.F.; Cross, F.T.

    1992-10-01

    Chromosome aberration frequency provides the most reliable biological marker of dose for detecting acute accidental radiation exposure. Significant radiation-induced changes in the frequency of chromosome aberrations can be detected at very low doses. Our paper provides information on using molecular chromosome probes paints'' to score chromosome damage and illustrates how technical advances make it possible to understand mechanisms involved during formation of chromosome aberrations. In animal studies chromosome aberrations provide a method to relate cellular damage to cellular dose. Using an In vivo/In vitro approach aberrations provided a biological marker of dose from radon progeny exposure which was used to convert WLM to dose in rat tracheal epithelial cells. Injection of Chinese hamsters with [sup 144]Ce which produced a low dose rate exposure of bone marrow to either low-LET radiation increased the sensitivity of the cells to subsequent external exposure to [sup 60]Co. These studies demonstrated the usefulness of chromosome damage as a biological marker of dose and cellular responsiveness.

  11. Metaphase chromosome aberrations as markers of radiation exposure and dose

    Energy Technology Data Exchange (ETDEWEB)

    Brooks, A.L.; Khan, M.A.; Jostes, R.F.; Cross, F.T.

    1992-10-01

    Chromosome aberration frequency provides the most reliable biological marker of dose for detecting acute accidental radiation exposure. Significant radiation-induced changes in the frequency of chromosome aberrations can be detected at very low doses. Our paper provides information on using molecular chromosome probes ``paints`` to score chromosome damage and illustrates how technical advances make it possible to understand mechanisms involved during formation of chromosome aberrations. In animal studies chromosome aberrations provide a method to relate cellular damage to cellular dose. Using an In vivo/In vitro approach aberrations provided a biological marker of dose from radon progeny exposure which was used to convert WLM to dose in rat tracheal epithelial cells. Injection of Chinese hamsters with {sup 144}Ce which produced a low dose rate exposure of bone marrow to either low-LET radiation increased the sensitivity of the cells to subsequent external exposure to {sup 60}Co. These studies demonstrated the usefulness of chromosome damage as a biological marker of dose and cellular responsiveness.

  12. Amphitelic orientation of centromeres at metaphase I is an important ...

    Indian Academy of Sciences (India)

    2014-07-31

    Jul 31, 2014 ... nick translation following the manufacturer's instructions. Chromosome observations were made and documented using an Olympus BX-51 microscope coupled to a Photometric. SenSys Olympus DP70 CCD camera (Tokyo, Japan). Results. Synthetic hexaploid wheat line Syn-SAU-6 was successfully.

  13. Metaphase chromosome aberrations as markers of radiation exposure and dose

    International Nuclear Information System (INIS)

    Brooks, A.L.; Khan, M.A.; Jostes, R.F.; Cross, F.T.

    1992-10-01

    Chromosome aberration frequency provides the most reliable biological marker of dose for detecting acute accidental radiation exposure. Significant radiation-induced changes in the frequency of chromosome aberrations can be detected at very low doses. Our paper provides information on using molecular chromosome probes ''paints'' to score chromosome damage and illustrates how technical advances make it possible to understand mechanisms involved during formation of chromosome aberrations. In animal studies chromosome aberrations provide a method to relate cellular damage to cellular dose. Using an In vivo/In vitro approach aberrations provided a biological marker of dose from radon progeny exposure which was used to convert WLM to dose in rat tracheal epithelial cells. Injection of Chinese hamsters with 144 Ce which produced a low dose rate exposure of bone marrow to either low-LET radiation increased the sensitivity of the cells to subsequent external exposure to 60 Co. These studies demonstrated the usefulness of chromosome damage as a biological marker of dose and cellular responsiveness

  14. NACK kinesin is required for metaphase chromosome alignment and cytokinesis in the moss Physcomitrella patens.

    Science.gov (United States)

    Naito, Haruko; Goshima, Gohta

    2015-01-01

    The NACK kinesins (NACK1, NACK2 in tobacco and AtNACK1/HINKEL, AtNACK2/STUD/TETRASPORE in Arabidopsis), members of a plant-specific kinesin-7 family, are required for cytokinesis. Previous studies using tobacco and Arabidopsis cells showed that NACK1 and AtNACK1 at the phragmoplast midzone activate the MAP kinase cascade during the late M phase, which is critical for the cell plate formation. However, the loss-of-function phenotype has not been investigated in details in living cells and the molecular activity of this kinesin remains to be determined. Here, we report the mitotic roles and activity of the NACK kinesins in the moss Physcomitrella patens. When we simultaneously knocked down three PpNACKs by RNA-interference (RNAi) in protonemal cells, we observed a cytokinesis failure following a defect in phragmoplast expansion. In addition, misaligned chromosomes were frequently detected in the pre-anaphase spindle and the anaphase onset was significantly delayed, indicating that PpNACK also plays a role in pre-anaphase. Consistent with the appearance of early and late mitotic phenotypes, endogenous PpNACK was localised to the interpolar microtubule (MT) overlap from prometaphase through telophase. In vitro MT gliding assay and single motor motility assay showed that PpNACK-b is a processive, plus-end-directed motor, suggesting that PpNACK is capable of transporting cargoes along the spindle/phragmoplast MT. Our study using Physcomitrella patens demonstrated that PpNACK is an active motor protein and identified unexpected and conserved roles of PpNACK during the mitosis of P. patens.

  15. Modified Method of Metaphase Plates Obtaining for Polyploid Fish Genera Carassius and Cobitis Karyotyping (Actinopterygii, Cypriniformes

    Directory of Open Access Journals (Sweden)

    Pukhtayevych P. P.

    2014-08-01

    Full Text Available Ото- браны и апробированы современные методики получения метафазных пластинок соматических клеток рыб, дающих наибольший эффект при исследовании полиплоидных видов. На основании эмпирических данных в отношении представителей родов Carassius и Cobitis рекомендуется ме- тодика с использованием CoCl2 и колхицина. В статье приведено подробное описание модифи- цированной методики и полученные в результате её применения метафазные пластинки Carassius auratus Linnaeus, 1758, Carassius carassius Linnaeus, 1758 и триплоидной формы Cobitis taenia Linnaeus, 1758.

  16. Diagnostic yield by supplementing prenatal metaphase karyotyping with MLPA for microdeletion syndromes and subtelomere imbalances

    DEFF Research Database (Denmark)

    Kjaergaard, S; Sundberg, K; Jørgensen, F S

    2010-01-01

    The aim of the study was to retrospectively assess the relevance of using multiplex ligation-dependent probe amplification (MLPA) for detection of selected microdeletion syndromes (22q11, Prader-Willi/Angelman, Miller-Dieker, Smith-Magenis, 1p-, Williams), the reciprocal microduplication syndrome...

  17. Proteomic analysis of human metaphase chromosomes reveals Topoisomerase II alpha as an Aurora B substrate

    DEFF Research Database (Denmark)

    Morrison, Ciaran; Henzing, Alexander J; Jensen, Ole Nørregaard

    2002-01-01

    The essential Aurora B kinase is a chromosomal passenger protein that is required for mitotic chromosome alignment and segregation. Aurora B function is dependent on the chromosome passenger, INCENP. INCENP, in turn, requires sister chromatid cohesion for its appropriate behaviour. Relatively few...... composition of the extracted chromosome fraction. Cloning, fluorescent tagging and expression in HeLa cells of the putative GTP-binding protein NGB/CRFG demonstrated it to be a novel mitotic chromosome protein, with a perichromosomal localisation. Identi fication of the protein bands corresponding to those...

  18. Diagnostic yield by supplementing prenatal metaphase karyotyping with MLPA for microdeletion syndromes and subtelomere imbalances

    DEFF Research Database (Denmark)

    Kjaergaard, S; Sundberg, K; Jørgensen, F S

    2010-01-01

    The aim of the study was to retrospectively assess the relevance of using multiplex ligation-dependent probe amplification (MLPA) for detection of selected microdeletion syndromes (22q11, Prader-Willi/Angelman, Miller-Dieker, Smith-Magenis, 1p-, Williams), the reciprocal microduplication syndromes...

  19. THE 3-GROUP METAPHASE AS A MORPHOLOGIC INDICATOR OF HIGH-PLOIDY CELLS IN CERVICAL INTRAEPITHELIAL NEOPLASIA

    NARCIS (Netherlands)

    PIETERS, WJLM; KOUDSTAAL, J; PLOEMZAAYER, JJ; Janssens, J; OOSTERHUIS, JW

    There is a need for additional morphologic criteria to improve the value of histologic classification for the prediction of the biologic behavior of cervical intraepithelial neoplasia (CIN). Representative slides from 72 cone specimens containing CIN were examined to study the correlation between

  20. Prophase I arrest and progression to metaphase I in mouse oocytes: comparison of resumption of meiosis and recovery from G2-arrest in somatic cells

    Czech Academy of Sciences Publication Activity Database

    Šolc, Petr; Schultz, R. M.; Motlík, Jan

    2010-01-01

    Roč. 16, č. 9 (2010), s. 654-664 ISSN 1360-9947 R&D Projects: GA ČR(CZ) GC301/09/J036; GA MŠk ME08030 Grant - others:NIH(US) HD22681 Institutional research plan: CEZ:AV0Z50450515 Keywords : resumption of meiosis * prophase I arrest * oocyte Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.506, year: 2010

  1. Development of porcine embryos reconstituted with somatic cells and enucleated metaphase I and II oocytes matured in a protein-free medium

    Directory of Open Access Journals (Sweden)

    Gibbons John R

    2001-07-01

    Full Text Available Abstract Background Many cloned animals have been created by transfer of differentiated cells at G0/G1 or M phase of the cell cycle into enucleated M II oocytes having high maturation/meiosis/mitosis-promoting factor activity. Because maturation/meiosis/mitosis-promoting factor activity during oocyte maturation is maximal at both M I and M II, M I oocytes may reprogram differentiated cell nuclei as well. The present study was conducted to examine the developmental ability in vitro of porcine embryos reconstructed by transferring somatic cells (ear fibroblasts into enucleated M I or M II oocytes. Results Analysis of the cell cycle stages revealed that 91.2 ± 0.2% of confluent cells were at the G0/G1 phase and 54.1 ± 4.4% of nocodazole-treated cells were at the G2/M phase, respectively. At 6 h after activation, nuclear swelling was observed in 50.0-88.9% and 34.4-39.5% of embryos reconstituted with confluent cells and nocodazole-treated cells regardless of the recipient oocytes, respectively. The incidence of both a swollen nucleus and polar body was low (6.3-10.5% for all nocodazole-treated donor cell regardless of the recipient oocyte. When embryos reconstituted with confluent cells and M I oocytes were cultured, 2 (1.5% blastocysts were obtained and this was significantly (P Conclusions Porcine M I oocytes have a potential to develop into blastocysts after nuclear transfer of somatic cells.

  2. A note on chromosomes of Pontellopsis herdmani and Pontella princeps (Copepoda) from the Laccadive sea

    Digital Repository Service at National Institute of Oceanography (India)

    Goswami, U.; Goswami, S.C.

    Pontellopsis herdmani and Pontella princeps (Pontellidae, Calanoida, Copepoda) showed a diploid number of 20 and a haploid number of 10 chromosomes during the spermatogonial metaphase and metaphase II stages. The chromosomes were in the size range...

  3. 40 CFR 79.65 - In vivo sister chromatid exchange assay.

    Science.gov (United States)

    2010-07-01

    ... metaphase portion of the cell cycle and presumably requires the enzymatic incision, translocation and... to optimize the sample collection time for second division metaphase cells. (ii) This assay may be... added to arrest cell growth in c-metaphase. Cells are harvested 4 hours later and second-division...

  4. Human oocyte chromosome analysis: complicated cases and major ...

    Indian Academy of Sciences (India)

    trast to mitotic cell cultures, the assessment of chromosomal. Figure 7. Prematurely condensed sperm chromatin in uncleaved oocytes. (a) Part of metaphase with 12 oocyte chromosomes (num- bered) and sperm chromatids of which the smaller ones (arrows) re- semble oocyte chromatids. (b) Part of metaphase with single ...

  5. Analysis of chromosome rearrangements on the basis of synaptonemal complexes in the offspring of mice exposed to γ-rays

    International Nuclear Information System (INIS)

    Kalikinskaya, E.I.; Bogdanov, Yu.F.; Kolomiets, O.L.; Shevchenko, V.A.

    1986-01-01

    Electron-microscopic analysis of synaptonemic complexes (SC), spread on the hypophase surface, was conducted to investigate chromosome rearrangements in sterile and semisterile F 1 malemause offsprings, exposed to 5 Gy γ-rays Paralelly Chromosome rearrangement account in diakinesis-metaphase 1 was conducted using light microscope, in the same animals. During SC analysis in pachytene chromosome rearrangements were found in 63% of spermatocytes. Under chromosome analysis in diakinesis-metaphase 1 in the same animals chromosome rearrangements were found only in 32% of cells. SC analysis allows one to reveal chromosome rearrangements, which can not be revealed in diakinesis-metaphase 1

  6. Prenatal diagnosis and molecular cytogenetic characterization of a 1.07-Mb microdeletion at 5q35.2–q35.3 associated with NSD1 haploinsufficiency and Sotos syndrome

    Directory of Open Access Journals (Sweden)

    Chih-Ping Chen

    2014-12-01

    Conclusion: Fetuses with Sotos syndrome may present macrocephaly, polyhydramnios, ventriculomegaly, and pyelectasis in the third trimester. aCGH and metaphase FISH are useful for rapid diagnosis of 5q35 microdeletion associated with Sotos syndrome.

  7. The Y chromosome of the Atelidae family (Platyrrhini): study by chromosome microdissection.

    Science.gov (United States)

    Gifalli-Iughetti, C; Koiffmann, C P

    2009-01-01

    In order to study the intergeneric variability of the Y chromosome, we describe the hybridization of the Y chromosome of Brachytelesarachnoides, obtained by microdissection, to metaphases of Atelesbelzebuthmarginatus, Lagothrixlagothricha, and Alouatta male specimens. Brachytelesarachnoides (Atelinae) has 62 chromosomes and a very small Y chromosome. Our results showed that the Brachytelesarachnoides Y chromosome probe hybridized to Lagothrixlagothricha metaphases yielding one hybridization signal on only the tiny Y chromosome, and when hybridized with Atelesbelzebuthmarginatus metaphases it yielded one hybridization signal on two thirds of the small acrocentric Y chromosome. However, no hybridization signal was observed in Alouatta metaphases (subfamily Alouattinae), a closely related genus in the Atelidae family. Furthermore, our data support a close phylogenetic relationship among Brachyteles, Ateles, and Lagothrix and their placement in the Atelinae subfamily, but exclude Alouatta from this group indicating its placement as basal to this group. Copyright 2009 S. Karger AG, Basel.

  8. Journal of Genetics | Indian Academy of Sciences

    Indian Academy of Sciences (India)

    GGAT)4, (GT)7 and (TA)10 onto metaphase chromosomes of the Pacific oyster, Crassostrea gigas. Hybridization signals were observed in all the SSR probes, but the distribution and intensity of signals varied according to the oligonucleotide repeat ...

  9. The human neurofilament gene (NEFL) is located on the short arm of chromosome 8.

    NARCIS (Netherlands)

    J. Hurst; D. Flavell (David); J-P. Julien (Jean-Pierre); D.N. Meijer (Dies); W. Mushynski (Walter); F.G. Grosveld (Frank)

    1987-01-01

    textabstractWe have localized the gene coding for the human neurofilament light chain (NEFL) to chromosome band 8p2.1 by Southern blotting of DNA from hybrid cell panels and in situ hybridization to metaphase chromosomes.

  10. Biological dosimetry in the case of large-scale accidental overexposure to ionising radiation: contribution of the automatic detection of dicentrics

    International Nuclear Information System (INIS)

    Vaurijoux, A.; Gruel, G.; Roch-Lefevre, S.; Gregoire, E.; Martin, C.; Voisin, P.; Voisin, P.H.; Roy, L.

    2010-01-01

    Following large-scale accidental overexposure to ionising radiation, a rapid triage of the exposed population can be performed by scoring dicentrics and ring chromosomes in 50 metaphases. This is rapid but not accurate. Following this triage step, the dose can be estimated by scoring 500 metaphases. This is lengthy but very accurate. To improve the methodology, we propose using software for automatic dicentric scoring, which was tested on 63 victims of a radiological accident. The method using automatic dicentric scoring was compared with the standard methods of manual scoring. The comparison of the doses obtained shows that the automatic method could replace the two steps of the manual scoring (50 and 500 metaphases). In fact, the automatic scoring method is more accurate than the manual scoring of 50 metaphases and can therefore be used for triage. (authors)

  11. Biological dosimetry in the case of large-scale accidental overexposure to ionising radiation: contribution of the automatic detection of dicentrics; Dosimetrie biologique en cas d'exposition radiologique d'un grand nombre d'individus: apport de la detection automatique des dicentriques

    Energy Technology Data Exchange (ETDEWEB)

    Vaurijoux, A.; Gruel, G.; Roch-Lefevre, S.; Gregoire, E.; Martin, C.; Voisin, P.; Voisin, P.H.; Roy, L. [Laboratoire de Dosimetrie Biologique, IRSN, 92 - Fontenay-aux-Roses (France)

    2010-04-15

    Following large-scale accidental overexposure to ionising radiation, a rapid triage of the exposed population can be performed by scoring dicentrics and ring chromosomes in 50 metaphases. This is rapid but not accurate. Following this triage step, the dose can be estimated by scoring 500 metaphases. This is lengthy but very accurate. To improve the methodology, we propose using software for automatic dicentric scoring, which was tested on 63 victims of a radiological accident. The method using automatic dicentric scoring was compared with the standard methods of manual scoring. The comparison of the doses obtained shows that the automatic method could replace the two steps of the manual scoring (50 and 500 metaphases). In fact, the automatic scoring method is more accurate than the manual scoring of 50 metaphases and can therefore be used for triage. (authors)

  12. Intravenous but not intragastric urogastrone-EGF is trophic to the intestine of parenterally fed rats.

    OpenAIRE

    Goodlad, R A; Wilson, T J; Lenton, W; Gregory, H; McCullagh, K G; Wright, N A

    1987-01-01

    The effects of beta-urogastrone/human epidermal growth factor (URO-EGF) on intestinal epithelial cell proliferation were studied in rats in which intestinal cell proliferation had been reduced to a steady state basal level, by maintaining the rats on total parenteral nutrition. The accumulation of arrested metaphases over a two hour time period was determined in a dose response study. Increasing doses of URO-EGF progressively raised the two hour collection of metaphases and intestinal weights...

  13. Meiotic behaviour and spermatogenesis in male mice heterozygous for translocation types also occurring in man

    International Nuclear Information System (INIS)

    Nijhoff, J.H.

    1981-01-01

    In this thesis a start was made with meiotic observations of mouse translocation types - a Robertsonian translocation and a translocation between a metacentric and an acrocentric chromosome - which also occur in man. As an exogeneous factor of possible influence, the meiotic effects of two types of radiation (fission neutrons and X-rays) administered at relatively low doses 2 and 3 hours before prometaphase-metaphase II (probably during metaphase-anaphase I), were determined in Rb4Bnr/+-males. (Auth.)

  14. Cytological evidence of chromosomal rearrangement in the second meiotic division after exposure to X-rays

    International Nuclear Information System (INIS)

    Szemere, G.

    1982-01-01

    Metaphase II cells with unequal dyad-arms and obvious X/autosomal rearrangements were found after an exposure to X-rays (2 Gy) of male mice at different stages of meiosis (pachytene, diplotene and diakinesis) with a frequency of 0.2, 1.26 and 0.6%, respectively, giving a direct cytological evidence of structural chromosomal rearrangements in metaphase II cells, partly with autosomal and partly with X/autosomal partners. (author)

  15. Radiation-induced chromosomal aberrations in the lymphocytes of various species of mammals and the influence of coffeine during the G-2 phase

    International Nuclear Information System (INIS)

    Rosenthal, M.

    1983-01-01

    The cellular kinetics and the G0-radiation sensitivity of human, chimpanzee, swine and rabbit lymphocytes were investigated using the lymphocytes test system (Ham's F-10 Medium, PHA). Due to the integration of BrdU in the DNA (S-phase), the author was able to distinguish between first, second and third mitoses (M1, M2, M3) in accordance with the differential colouring of the metaphase chromosomes which took place according to the labelling pattern. When checking the G0-radiation sensitivity of the lymphocytes, the rates of chromosomal aberrations in the metaphases of the first and second mitoses were evaluated separately. The different radiation sensitivities are thought to be due to interspecies differences in the repair capacity of the lymphocytes. In the metaphases of second mitoses, the rate of dicentric chromosomes is approximately half of that in M1-metaphases. Ring chromosomes were nearly as frequent in M2-metaphases as in M1-metaphases. In the second experimental phase, the effects of coffein on the aberration rates after radiation exposure of the lymphocytes in the G2 phase was investigated. Achromatic lesions, open chromatide breaks, and translocations were evaluated. Aberration rates were found to increase with the radiation dose and to decrease with the cultivation time after radiation exposure. There was no marked effect of coffein on the aberration rates. The progress of the G2 phase was measured in terms of the rate of radioactively labelled metaphases, which increased with the cultivation time. This labelling index was lower in the exposed cultures than in the control cultures, suggesting a radiation-induced delay of the G2 phase. The labelling indexes of all cultures were enhanced after coffein treatment, suggesting a coffein-induced acceleration of the G2 phase. (orig./MG) [de

  16. Chromosomal instability in mouse embryonic fibroblasts null for the transcriptional co-repressor Ski

    Science.gov (United States)

    Marcelain, Katherine; Armisen, Ricardo; Aguirre, Adam; Ueki, Nobuhide; Toro, Jessica; Colmenares, Clemencia; Hayman, Michael J

    2011-01-01

    Ski is a transcriptional regulator that has been considered an oncoprotein, given its ability to induce oncogenic transformation in avian model systems. However, studies in mouse and in some human tumor cells have also indicated a tumor suppressor activity for this protein. We found that Ski−/− mouse embryo fibroblasts exhibit high levels of genome instability, namely aneuploidy, consistent with a tumor suppressor function for Ski. Time-lapse microscopy revealed lagging chromosomes and chromatin/chromosome bridges as the major cause of micronuclei formation and the subsequent aneuploidy. Although these cells arrested in mitosis after treatment with spindle disrupting drugs and exhibited a delayed metaphase/anaphase transition, Spindle Assembly Checkpoint (SAC) was not sufficient to prevent chromosome missegregation, consistent with a weakened SAC. Our in vivo analysis also showed dynamic metaphase plate rearrangements with switches in polarity in cells arrested in metaphase. Importantly, after ectopic expression of Ski the cells that displayed this metaphase arrest died directly during metaphase or after aberrant cell division, relating SAC activation and mitotic cell death. This increased susceptibility to undergo mitosis-associated cell death reduced the number of micronuclei-containing cells. The presented data support a new role for Ski in the mitotic process and in maintenance of genetic stability, providing insights into the mechanism of tumor suppression mediated by this protein. PMID:21412778

  17. Somatic pairing, endomitosis and chromosome aberrations in snakes (Viperidae and Colubridae

    Directory of Open Access Journals (Sweden)

    Beçak Maria Luiza

    2003-01-01

    Full Text Available The positioning of macrochromosomes of Bothrops jararaca and Bothrops insularis (Viperidae was studied in undistorted radial metaphases of uncultured cells (spermatogonia and oogonia not subjected to spindle inhibitors. Colchicinized metaphases from uncultured (spleen and intestine and cultured tissues (blood were also analyzed. We report two antagonic non-random chromosome arrangements in untreated premeiotic cells: the parallel configuration with homologue chromosomes associated side by side in the metaphase plate and the antiparallel configuration having homologue chromosomes with antipolar distribution in the metaphase ring. The antiparallel aspect also appeared in colchicinized cells. The spatial chromosome arrangement in both configurations is groupal size-dependent and maintained through meiosis. We also describe, in untreated gonia cells, endomitosis followed by reductional mitosis which restores the diploid number. In B. jararaca males we observed that some gonad regions present changes in the meiotic mechanism. In this case, endoreduplicated cells segregate the diplochromosomes to opposite poles forming directly endoreduplicated second metaphases of meiosis with the suppression of first meiosis. By a successive division, these cells form nuclei with one set of chromosomes. Chromosome doubling in oogonia is known in hybrid species and in parthenogenetic salamanders and lizards. This species also presented chromosome rearrangements leading to aneuploidies in mitosis and meiosis. It is suggested that somatic pairing, endomitosis, meiotic alterations, and chromosomal aberrations can be correlated processes. Similar aspects of nuclei configurations, endomitosis and reductional mitosis were found in other Viperidae and Colubridae species.

  18. In vivo FRET imaging revealed a regulatory role of RanGTP in kinetochore-microtubule attachments via Aurora B kinase.

    Directory of Open Access Journals (Sweden)

    Yoke-Peng Lee

    Full Text Available Under the fluctuating circumstances provided by the innate dynamics of microtubules and opposing tensions resulted from microtubule-associated motors, it is vital to ensure stable kinetochore-microtubule attachments for accurate segregation. However, a comprehensive understanding of how this regulation is mechanistically achieved remains elusive. Using our newly designed live cell FRET time-lapse imaging, we found that post-metaphase RanGTP is crucial in the maintenance of stable kinetochore-microtubule attachments by regulating Aurora B kinase via the NES-bearing Mst1. More importantly, our study demonstrates that by ensuring stable alignment of metaphase chromosomes prior to segregation, RanGTP is indispensible in governing the genomic integrity and the fidelity of cell cycle progression. Our findings suggest an additional role of RanGTP beyond its known function in mitotic spindle assembly during the prometaphase-metaphase transition.

  19. Chromosomal aberrations induced in mice bone marrow by treating with cisplatin and irradiation

    International Nuclear Information System (INIS)

    Wuerschmidt, F.; Molls, M.; Bardenheuer, M.J.; Mueller, W.U.

    2000-01-01

    Background: The aim of this study was to quantify the combined effect of cisplatin and radiation on chromosomal damage with emphasis on the time interval between cisplatin and radiation. Methods and materials: Bone marrow of female NMRI-nu(+)mice was taken as a model system for a highly proliferative tissue irradiated with cobalt-60 (1 to 4 Gy). Cisplatin was injected intraperitoneally (ip) at 1.1 to 36 mg/kg. Cisplatin was given at various time intervals before and after radiation. Bone marrow and metaphases were prepared according to standard procedures. Results: The percentage of aberrant metaphases after radiation or cisplatin alone increased in a dose-dependent manner (sigmoidal dose-response curve). Combining both modalities led to additive values at all time points for the percentage of aberrant metaphases. Borderline significant (p [de

  20. Mitosis-specific phosphorylation of PML at T409 regulates spindle checkpoint.

    Science.gov (United States)

    Jin, J; Liu, J

    2016-08-31

    During mitosis, Promyelocytic leukemia nuclear bodies (PML NBs) change dramatically in morphology and composition, but little is known about function of PML in mitosis. Here, we show that PML is phosphorylated at T409 (PML p409) in a mitosis-specific manner. More importantly, PML p409 contributes to maintain the duration of pro-metaphase and regulates spindle checkpoint. Deficient PML p409 caused a shortening of pro-metaphase and challenged the nocodazole-triggered mitotic arrest. T409A mutation led to a higher frequency of misaligned chromosomes on metaphase plate, and subsequently death in late mitosis. In addition, inhibition of PML p409 repressed growth of tumor cells, suggesting that PML p409 is a potential target for cancer therapy. Collectively, our study demonstrated an important phosphorylated site of PML, which contributed to explore the role of PML in mitosis.

  1. Aurora A's functions during mitotic exit: the Guess Who game

    Directory of Open Access Journals (Sweden)

    David eReboutier

    2015-12-01

    Full Text Available Until recently, the knowledge of Aurora A kinase functions during mitosis was limited to pre-metaphase events, particularly centrosome maturation, G2/M transition, and mitotic spindle assembly. However, an involvement of Aurora A in post-metaphase events was also suspected, but not clearly demonstrated due to the technical difficulty to perform the appropriate experiments. Recent developments of both an analog specific version of Aurora A, and of small molecule inhibitors have led to the first demonstration that Aurora A is required for the early steps of cytokinesis. As in pre-metaphase, Aurora A plays diverse functions during anaphase, essentially participating in astral microtubules dynamics and central spindle assembly and functioning. The present review describes the experimental systems used to decipher new functions of Aurora A during late mitosis and situate these functions into the context of cytokinesis mechanisms.

  2. FISH on chromosomes derived from the snail model organism Biomphalaria glabrata.

    Science.gov (United States)

    Odoemelam, Edwin C; Raghavan, Nithya; Ittiprasert, Wannaporn; Miller, Andre; Bridger, Joanna M; Knight, Matty

    2010-01-01

    The application of fluorescence in situ hybridization (FISH) for the mapping of single copy genes onto homologous chromosome has been integral to vast number genome sequencing projects, such as that of mouse and human. The chromosomes of these organisms are well-studied and are the staple resource of most of the early studies conducted in cytogenetics. However, there are now protocols for analyzing FISH probes in a number of different organisms on both metaphase and interphase chromosomes.Here, we describe the methodologies for the chromosomal mapping of nonrepetitive (single-copy) genes of the snail Biomphalaria glabrata onto metaphase chromosomes derived from the only molluscan cell-line in existence. The technique described in this chapter was developed for the B. glabrata genome sequencing project through troubleshooting experimental procedures established for other organisms so that both the optimum resolution of metaphase chromosome and the effective hybridization of genes were achieved.

  3. Cytogenetic evidence for genome elimination during microsporogenesis in interspecific hybrid between Brachiaria ruziziensis and B. brizantha (Poaceae

    Directory of Open Access Journals (Sweden)

    Andréa Beatriz Mendes-Bonato

    2006-01-01

    Full Text Available Microsporogenesis was analyzed in an interspecific hybrid between an artificially tetraploidized sexual accession of Brachiaria ruziziensis (R genome and a natural apomictic tetraploid accession of B. brizantha (B genome. Chromosomes associated predominantly as bivalents. From this phase to the end of meiosis, chromosomes presented irregular segregation and abnormal arrangement in the metaphase plate. During metaphase I, in 27.8% of meiocytes, bivalents were distributed in two metaphase plates. In anaphase I, two distinct and typical bipolar spindles were formed. In 29.7% of pollen mother cells, one genome did not divide synchronically, with chromosomes lagging behind or not segregating at all. The second division was very irregular, resulting in polyads. Based on previous results from analysis of a triploid hybrid between these species, where the R genome was eliminated by asynchrony during meiosis, it is suggested that the laggard genome in this hybrid also belongs to B. ruziziensis.

  4. Chromosome number distribution and cellular DNA content in colorectal adenomas from polyposis and nonpolyposis patients

    DEFF Research Database (Denmark)

    Petersen, S E; Madsen, A L; Bak, Martin

    1991-01-01

    and a correspondingly increased nuclear DNA content. In another two adenomas, the DNA analyses showed small hyperploid populations constituting 6% and 2% of the cells. The most striking difference between the DNA analyses and chromosome number distributions was that 13% of all metaphases were hyperploid with chromosome......Ploidy analyses of colorectal adenomas were performed by combined flow cytometric DNA analysis of unfixed isolated nuclei and direct chromosome preparation after Colcemid incubation for 9-20 hours. Ten of 18 adenomas from nonpolyposis patients and 4 of 13 adenomas from patients with familial...... adenomatous polyposis yielded a mean of 25 countable metaphases (range 7-44) per tumor. Of 343 metaphases, only 38% had 46 chromosomes, and 62% were nondiploid. All but one adenoma had diploid or peridiploid modes in the range of 46-50 chromosomes. One adenoma was hyperploid, with a mode of 74 chromosomes...

  5. Procedure Improvement in Blood Processing for Chromosome Aberration Analyst

    International Nuclear Information System (INIS)

    Noraisyah Mohd Yusof; Juliana Mahamad; Rahimah Abd Rahim; Yahaya Talib; Mohd Rodzi Ali

    2015-01-01

    Detection of chromosome at metaphase of the cell cycle is performed either manually or automatically. Procedure for slide preparation published by the IAEA does not guarantee that the quality of slide is suitable for automatic detection. The detection efficiency reduces if there is cells debris on slides. This paper describes the modifications made to the standard procedure. The period of hypotonic treatment to the cell was lengthened; the slides were pre-treated with RNase and the frequency of rinsing during the chromosomal coloring process was increased. Results show the metaphase images were better and clearer, and numbers of metaphase that can be detected automatically were also increased. In conclusion, modification to the current standard protocol helps to easy the process of chromosome aberration analysis at Nuclear Malaysia. (author)

  6. DNA damage response and spindle assembly checkpoint function throughout the cell cycle to ensure genomic integrity.

    Directory of Open Access Journals (Sweden)

    Katherine S Lawrence

    2015-04-01

    Full Text Available Errors in replication or segregation lead to DNA damage, mutations, and aneuploidies. Consequently, cells monitor these events and delay progression through the cell cycle so repair precedes division. The DNA damage response (DDR, which monitors DNA integrity, and the spindle assembly checkpoint (SAC, which responds to defects in spindle attachment/tension during metaphase of mitosis and meiosis, are critical for preventing genome instability. Here we show that the DDR and SAC function together throughout the cell cycle to ensure genome integrity in C. elegans germ cells. Metaphase defects result in enrichment of SAC and DDR components to chromatin, and both SAC and DDR are required for metaphase delays. During persistent metaphase arrest following establishment of bi-oriented chromosomes, stability of the metaphase plate is compromised in the absence of DDR kinases ATR or CHK1 or SAC components, MAD1/MAD2, suggesting SAC functions in metaphase beyond its interactions with APC activator CDC20. In response to DNA damage, MAD2 and the histone variant CENPA become enriched at the nuclear periphery in a DDR-dependent manner. Further, depletion of either MAD1 or CENPA results in loss of peripherally associated damaged DNA. In contrast to a SAC-insensitive CDC20 mutant, germ cells deficient for SAC or CENPA cannot efficiently repair DNA damage, suggesting that SAC mediates DNA repair through CENPA interactions with the nuclear periphery. We also show that replication perturbations result in relocalization of MAD1/MAD2 in human cells, suggesting that the role of SAC in DNA repair is conserved.

  7. α-endosulfine (ENSA) regulates exit from prophase I arrest in mouse oocytes

    OpenAIRE

    Matthews, Lauren M; Evans, Janice P

    2014-01-01

    Mammalian oocytes in ovarian follicles are arrested in meiosis at prophase I. This arrest is maintained until ovulation, upon which the oocyte exits from this arrest, progresses through meiosis I and to metaphase of meiosis II. The progression from prophase I to metaphase II, known as meiotic maturation, is mediated by signals that coordinate these transitions in the life of the oocyte. ENSA (α-endosulfine) and ARPP19 (cAMP-regulated phosphoprotein-19) have emerged as regulators of M-phase, w...

  8. Heavy ion-induced chromosomal aberrations analyzed by fluorescence in situ hybridization

    Energy Technology Data Exchange (ETDEWEB)

    Durante, M.; Gialanella, G.; Grossi, G.; Pugliese, M. [Univ. ``Federico II``, Naples (Italy). Dept. of Physics]|[INFN, Naples (Italy); Cella, L.; Greco, O. [Univ. ``Federico II``, Naples (Italy). Dept. of Physics; Furusawa, Y. [NIRS, Chiba (Japan); George, K.; Yang, T.C. [NASA Lyndon B. Johnson Space Center, Houston, TX (United States)

    1997-09-01

    We have investigated the effectiveness of heavy ions in the induction of chromosomal aberrations in mammalian cells by the recent technique of fluorescence in situ hybridization (FISH) with whole-chromosome probes. FISH-painting was used both in metaphase and interphase (prematurely condensed) chromosomes. The purpose of our experiments was to address the following problems: (a) the ratio of different types of aberrations as a function of radiation quality (search for biomarkers); (b) the ratio between aberrations scored in interphase and metaphase as a function of radiation quality (role of apoptosis); (c) differences between cytogenetic effects produced by different ions at the same LET (role of track structure). (orig./MG)

  9. Possible mechanisms of chromosomal aberrations: VII. Comparative dynamics of sister chromatid disjunction and realization of radiation-induced chromosomal aberrations during mitosis

    International Nuclear Information System (INIS)

    Lebedeva, L.I.; Akhmamet'eva, E.M.

    1994-01-01

    An increase in radiation-induced chromosomal aberrations during c-metaphase sister chromatid disjunction was demonstrated in murine bone marrow cells exposed to a total γ-irradiation at 0.5 Gy. Caffeine (Cf) treatment during mitosis partially suppressed the chromatid disjunction rate and increased the number of radiation-induced aberrations in this mitosis. Nalidixic acid (NA) treatment of c-metaphase cells completely suppressed chromatid disjunction and the realization of induced aberrations. Topoisomerase 2 was assumed to be involved during mitosis in both processes

  10. A Semi-Closed Device for Chromosome Spreading for Cytogenetic Analysis

    DEFF Research Database (Denmark)

    Kwasny, Dorota; Mednova, Olga; Vedarethinam, Indumathi

    2014-01-01

    Metaphase chromosome spreading is the most crucial step required for successful karyotyping and FISH analysis. These two techniques are routinely used in cytogenetics to assess the chromosome abnormalities. The spreading process has been studied for years but it is still considered an art more than......, and thus allow for better quality spreads. Further cytogenetic analysis depends on the quality of the spreads, which is dependent on the skills of the personnel and is thus limited to laboratory settings. Here, we present a semi-closed microfluidic chip for preparation of the metaphase spreads on a glass...

  11. Meiotic behaviour in three interspecific three-way hybrids between ...

    Indian Academy of Sciences (India)

    Chromosome paired predominantly as multivalents suggesting that genetic recombination and introgression of specific target genes from B. brizantha into B. ruziziensis can be expected. Arrangement of parental genomes in distinct metaphase plates was observed in H27 and H34, which have different male genitors.

  12. Karyotype and nucleic acid content in Zantedeschia aethiopica Spr ...

    African Journals Online (AJOL)

    Analysis of karyotype, nucleic deoxyribonucleic acid (DNA) content and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) were performed in Zantedeschia aethiopica and Zantedeschia elliottiana. Mitotic metaphase in both species showed 2n=32. The chromosomes of both species were quite similar ...

  13. Marked chromosomes associations in catarrhine monkeys, with a note on chromosome associations in other primate groups

    NARCIS (Netherlands)

    Boer, L.E.M. de

    In metaphase figures, obtained from cultures of whole blood, associations of the marked chromosomes were found in species of the following genera: Macaca, Cercocebus, Cercopithecus and Symphalangus. The different types of these associations are discussed. A note is given on chromosome associations

  14. Mutagenic Potentials of the Sterilizing Fluid – Puritil on Root Tip ...

    African Journals Online (AJOL)

    The sterilizing fluid was found to exhibit both mitoclassic and chromatoclassic effects and thus induced a number of abnormalities. These abnormalities included disturbed interphase involving nuclear lesions and nuclear dissolutions, micronuclei and binucleate cells, sticky metaphase, disturbed anaphase involving ...

  15. Completion of meiosis in male zebrafish (Danio rerio) despite lack of DNA mismatch repair gene mlh1.

    NARCIS (Netherlands)

    Leal, M.C.; Feitsma, H.; Cuppen, E.; Franca, L.R.; Schulz, R.W.

    2008-01-01

    Mlh1 is a member of DNA mismatch repair (MMR) machinery and is also essential for the stabilization of crossovers during the first meiotic division. Recently, we have shown that zebrafish mlh1 mutant males are completely infertile because of a block in metaphase I, whereas females are fertile but

  16. acetyl (Lys14)-histone H3 predominantly mark the pericentromeric ...

    Indian Academy of Sciences (India)

    2016-12-01

    Dec 1, 2016 ... chromosomes during meiotic metaphase I. Colocalization of anti-H3S10phK14ac with an onion's CENH3 antibody on mitotic chromosomes of Allium revealed the chromosomal location .... prophase stages in all the plants during mitosis (figure 1, a and i; figure 2a ; arrow marked). The immunosignals could.

  17. CLIP-170 facilitates the formation of kinetochore-microtubule attachments

    NARCIS (Netherlands)

    Tanenbaum, ME; Galjart, N; van Vugt, MATM; Medema, RH

    2006-01-01

    CLIP-170 is a microtubule 'plus end tracking' protein involved in several microtubule-dependent processes in interphase. At the onset of mitosis, CLIP-170 localizes to kinetochores, but at metaphase, it is no longer detectable at kinetochores. Although RNA interference (RNAi) experiments have

  18. 2092-IJBCS-Article-Olufemi Awodiran

    African Journals Online (AJOL)

    hp

    Metaphase chromosomes were freshly prepared from newly hatched larvae as described by Aluko and Awopetu (1995) though slightly modified by reducing the number of hours in Colchicine (BH15 1TD,. England) solution (Olaniyi, 2008). One hour old embryos of the fish were put in 0.01%. Colchicine solution for 3 h and ...

  19. 40 CFR 798.5375 - In vitro mammalian cytogenetics.

    Science.gov (United States)

    2010-07-01

    ... metaphase stages first appear in the culture will include cells in all phases of the division cycle... first post-treatment mitosis and numerical aberrations require at least one cell division to be... changes the cell cycle length, the fixation intervals should be changed accordingly. If a single harvest...

  20. Prenatal detection of aneuploidies using fluorescence in situ ...

    Indian Academy of Sciences (India)

    Unknown

    Fluorescence in situ hybridization (FISH) is a powerful molecular cytogenetic technique which allows rapid detection of aneuploidies on interphase cells and metaphase spreads. The aim of the present study was to evaluate FISH as a tool in prenatal diagnosis of aneuploidies in high risk pregnancies in an Indian set up.

  1. Stabilization of chromosomes by DNA intercalators for flow karyotyping and identification by banding of isolated chromosomes

    NARCIS (Netherlands)

    Aten, J. A.; Buys, C. H.; van der Veen, A. Y.; Mesa, J. R.; Yu, L. C.; Gray, J. W.; Osinga, J.; Stap, J.

    1987-01-01

    A number of structurally unrelated DNA intercalators have been studied as stabilizers of mitotic chromosomes during isolation from rodent and human metaphase cells. Seven out of the nine intercalators tested were found to be useful as chromosome stabilizing agents. Chromosome suspensions prepared in

  2. A rapid, non-sacrificial chromosome preparation technique for ...

    African Journals Online (AJOL)

    1987-09-16

    Sep 16, 1987 ... and view under oil. The range and quality of cells arrested in metaphase for both species was sufficient to permit karyotyping and chromosome counting. (See Figure 1a & Jb). The stress of hand!i ng the fish appeared to influence the mitotic index. This effect was reduced by painting the colchicine aquarium ...

  3. New trends and techniques in chromosome aberration analysis

    International Nuclear Information System (INIS)

    Bender, M.A.

    1978-01-01

    The following topics are discussed: automation of chromosome analysis; storage of fixed cells from cultures of lymphocytes obtained routinely during periodic employee medical examinations; analysis of banded chromosomes; identification of first division metaphases; sister chromatid exchange; and patterns of aberration induction

  4. QTL list: uni13.1 [PGDBj Registered plant list, Marker list, QTL list, Plant DB link and Genome analysis methods[Archive

    Lifescience Database Archive (English)

    Full Text Available QT61665 Brassica napus Brassicaceae uni13.1 controlling homeologous pairing at met...aphase I in haploids PrBn gene (homeologous pairing regulator in B. napus) ... LG_DY13 ... 43.4 2.5 ... 10.1534/genetics.106.064071 16951054

  5. QTL list: uni4.1 [PGDBj Registered plant list, Marker list, QTL list, Plant DB link and Genome analysis methods[Archive

    Lifescience Database Archive (English)

    Full Text Available QT61663 Brassica napus Brassicaceae uni4.1 controlling homeologous pairing at meta...phase I in haploids PrBn gene (homeologous pairing regulator in B. napus) ... LG_DY04 ... 135.67 3.3 ... 10.1534/genetics.106.064071 16951054

  6. QTL list: PrBn [PGDBj Registered plant list, Marker list, QTL list, Plant DB link and Genome analysis methods[Archive

    Lifescience Database Archive (English)

    Full Text Available QT61662 Brassica napus Brassicaceae PrBn controlling homeologous pairing at metaph...ase I in haploids PrBn gene (homeologous pairing regulator in B. napus) ... LG_DY15 ... 34.9 7.7 ... 10.1534/genetics.106.064071 16951054

  7. Karyotypes in Ethiopian Aloe species (Xanthorrhoeaceae: Asphodeloideae)

    DEFF Research Database (Denmark)

    Fentaw, Eshetu; Dagne, Kifle; Rønsted, Nina

    2013-01-01

    A cytogenetic survey of 18 species in the succulent-leaved genus Aloe L. (Xanthorrhoeaceae subfamily Asphodeloideae) in Ethiopia was undertaken towards a more complete genetic characterisation of the genus throughout its distribution in Africa, Arabia and Madagascar. Somatic metaphase chromosomes...

  8. Download this PDF file

    African Journals Online (AJOL)

    ADOWIE PERE

    Exonn Mobil floating production storage and off- loading facility while seeds of Arachis hypogaea .... sticky chromosomes, anaphase bridges and vagrant chromosome observed (plate 2). There were ... at (a) Prophase (b) metaphase (c) anaphase. (d) telophase (e) c-mitosis (f) sticky cell (g) anaphase bridge (h) vagrant. 5.7.

  9. The Detection and Analysis of Chromosome Fragile Sites

    DEFF Research Database (Denmark)

    Bjerregaard, Victoria A; Özer, Özgün; Hickson, Ian D

    2018-01-01

    A fragile site is a chromosomal locus that is prone to form a gap or constriction visible within a condensed metaphase chromosome, particularly following exposure of cells to DNA replication stress. Based on their frequency, fragile sites are classified as either common (CFSs; present in all...

  10. Molecular screening for Yq microdeletion in men with idiopathic ...

    Indian Academy of Sciences (India)

    Unknown

    forma. Peripheral blood cultures were setup for chromo- somal analysis in all the cases and 5 well spread G- banded metaphases were karyotyped using the ..... torchidism and AZF microdeletions; Asian J. Androl. 148. Dada R, Gupta N P and Kucheria K 2002b AZF microdeletions in cases with Cryptorchidism, Varicocele ...

  11. Insect chromosomes preparing methods for genetic researches ...

    African Journals Online (AJOL)

    Cytogenetics are almost always based on the examination of the fixed mitotic chromosomes during the analyses of metaphase. During this phase of the cycle of cells, the DNA is folded up and chromatin is strongly condensed. The relative position of the centromere is constant, which means that the ratio of the lengths of the ...

  12. The Use of Double-Monotelodisomics to Identify Translocations in Triticum aestivum

    DEFF Research Database (Denmark)

    Linde-Laursen, Ib; Larsen, J.

    1974-01-01

    By analysing metaphase I of double-monotelodisomic hybrids between two varieties of hexaploid wheat differentiated by reciprocal translocations it is possible to establish reliably the chromosomes involved in each translocation. Also the chromosome parts translocated may be identified. The use...

  13. Localization of introduced genes on the chromosomes of transgenic barley, wheat and triticale by fluorescence in situ hybridization

    DEFF Research Database (Denmark)

    Pedersen, C.; Zimny, J.; Becker, D.

    1997-01-01

    Using fluorescence in situ hybridization (FISH) we localized introduced genes on metaphase chromosomes of barley, wheat, and triticale transformed by microprojectile bombardment of microspores and scutellar tissue with the pDB1 plasmid containing the uidA and bar genes. Thirteen integration sites...

  14. Effect of cumulus-oocyte complexes (COCs) culture duration on in ...

    African Journals Online (AJOL)

    We investigated and optimized the cumulus-oocyte complexes (COCs) culture duration for pig oocyte in vitro maturation and produced a number of high-quality metaphase-II (M-II) oocytes for generation of parthenotes. The present study graded the COCs into levels A, B and C according to layers of cumulus cells, which ...

  15. culture duration on in vitro maturation and parthenogenetic ...

    African Journals Online (AJOL)

    user

    2010-12-23

    Dec 23, 2010 ... We investigated and optimized the cumulus-oocyte complexes (COCs) culture duration for pig oocyte in vitro maturation and produced a number of high-quality metaphase-II (M-II) oocytes for generation of parthenotes. The present study graded the COCs into levels A, B and C according to layers of ...

  16. Inhibiting Crm1 causes the formation of excess acentriolar spindle poles containing NuMA and B23, but does not affect centrosome numbers.

    Science.gov (United States)

    Rousselet, Annie

    2009-09-14

    B23/nucleophosmin is present on spindle poles at metaphase. Migration of B23 to the poles is under the control of exportin Crm1. B23 at the centrosome plays a role in the control centrosome duplication. h-Tert-RPE1 cells blocked in prometaphase with low doses of Nocodazol showed a progression to mitosis if Crm1 exportin was inhibited. Under these conditions, the formation of accessory poles containing gamma-tubulin, NuMA (nuclear-mitotic-apparatus) and B23 was induced at metaphase. No effect on centrosome number was observed. In quiescent h-Tert-RPE1 cells, when Crm1 was active, B23 was not detected at the centrosome as well as B23-mutants reported to block centrosome duplication. In addition, the modification of B23 nucleo-cytoplasmic shuttling showed no effect on centrosome duplication. Inhibition of Crm1 in early metaphase favours the formation of supplementary acentriolar spindle poles. B23 and NuMA are present at these poles that ultimately focus around the centrosome. Inhibition of Crm1 at metaphase has no effect on the control of centrosome numbers.

  17. Automictic parthenogenesis in the parasitoid Venturia canescens (Hymenoptera : Ichneumonidae) revisited

    NARCIS (Netherlands)

    Beukeboom, LW; Pijnacker, LP; Traut, W.

    2000-01-01

    Both arrhenotokous and thelytokous reproduction are known to occur in the parasitoid wasp Venturia canescens. The cytological mechanism of thelytoky was previously reported to involve the formation of a restitution metaphase after the reduction division, but the exact nature of the subsequent

  18. Time-optimized analysis of slit-scan chromosome profiles on a general-purpose personal computer

    NARCIS (Netherlands)

    Heilig, R.; Hausmann, M.; Rens, W.; Aten, J. A.; Cremer, C.

    1993-01-01

    Slit-scan flow cytometry provides a method to analyze large numbers of metaphase chromosomes in a relatively short time due to morphological features. The high detection rate requires fast computing for on-line analysis. Up to now, this has been achieved using special-purpose computers, parallel

  19. Somatic chromosomal abnormalities in couples undergoing infertility ...

    Indian Academy of Sciences (India)

    1999). Oligozoospermia is .... age of abnormal metaphases needed to distinguish between low level and true mosaicism. This borderline ... Moore A. K., Lynch K., Arny M. J. and Grow D. R. 2008 Turner mosaicism (45,X/46,XX) diagnosed in a ...

  20. A high resolution chromosome image processor for study purposes, NIRS-1000:CHROMO STUDY, and algorithm developing to classify radiation induced aberrations.

    Science.gov (United States)

    Yamamoto, M; Hayata, I; Furuta, S

    1992-03-01

    Since 1989 we have promoted a project to develop an automated scoring system of radiation induced chromosome aberrations. As a first step, a high resolution image processing system for study purposes, NIRS-1000:CHROMO STUDY, has been developed. It is composed of: (1) CHROMO MARKER whose main purpose is to mark on images to make image data base, (2) CHROMO ALGO whose purpose is algorithm development, and (3) METAPHASE RANKER whose purposes are metaphase finding and ranking with a high power objective lens. However, METAPHASE RANKER is presently under development. The system utilizes a high definition video system so as to realize the best spatial resolution that is achievable with an optical microscope using an objective lens (x 100, numerical aperture 1.4). The video camera has 1024 effective scan lines to realize 0.1 microns sampling on a specimen. The system resolution achieved on the hard copy is less than 0.3 microns on a specimen. A preliminary algorithm has been developed to classify the aberrations on the system using projection information of gray level. The preliminary test results on excellent 10 metaphases show that the correct classification ratio is 92.7%, that the detection rate of the aberrations is 83.3% and that the false positive rate is 6.1%.

  1. Supernumerary ring chromosome 20 characterized by fluorescence in situ hybridization

    NARCIS (Netherlands)

    Van Langen, Irene M.; Otter, Mariëlle A.; Aronson, Daniël C.; Overweg-Plandsoen, W.C.G.; Hennekam, Raoul C.M.; Leschot, Nico J.; Hoovers, Jan M.N.

    1996-01-01

    We report on a boy with mild dysmorphic features and developmental delay, in whom karyotyping showed an additional minute ring chromosome in 60% of metaphases. Fluorescence in situ hybridization (FISH) with a centromere specific probe demonstrated that the ring chromosome contained the centromeric

  2. Analysis of Chromothripsis by Combined FISH and Microarray Analysis.

    Science.gov (United States)

    MacKinnon, Ruth N

    2018-01-01

    Fluorescence in situ hybridization (FISH) to metaphase chromosomes, in conjunction with SNP array, array CGH, or whole genome sequencing, can help determine the organization of abnormal genomes after chromothripsis and other types of complex genome rearrangement. DNA microarrays can identify the changes in copy number, but they do not give information on the organization of the abnormal chromosomes, balanced rearrangements, or abnormalities of the centromeres and other regions comprised of highly repetitive DNA. Many of these details can be determined by the strategic use of metaphase FISH. FISH is a single-cell technique, so it can identify low-frequency chromosome abnormalities, and it can determine which chromosome abnormalities occur in the same or different clonal populations. These are important considerations in cancer. Metaphase chromosomes are intact, so information about abnormalities of the chromosome homologues is preserved. Here we describe strategies for working out the organization of highly rearranged genomes by combining SNP array data with various metaphase FISH methods. This approach can also be used to address some of the uncertainties arising from whole genome or mate-pair sequencing data.

  3. Journal of Genetics | Indian Academy of Sciences

    Indian Academy of Sciences (India)

    Studies on meiosis in pollen mother cells (PMCs) of a triploid interspecific hybrid ( 3 x = 39 chromosomes, AAD) between tetraploid Gossypium hirsutum ( 4 n = 2 x = 52 ,AADD) and diploid G. arboreum ( 2 n = 2 x = 26 ,AA) are reported. During meiotic metaphase I, 13 AA bivalents and 13 D univalents are expected in the ...

  4. Prenatal detection of aneuploidies using fluorescence in situ ...

    Indian Academy of Sciences (India)

    Multicolour commercially available FISH probes specific for chromosomes 13, 18, 21, X and Y were used. Interphase FISH was done on uncultured cells from chorionic villus and amniotic fluid samples. FISH on metaphase spreads was done from cord blood samples. The results of FISH were in conformity with the results of ...

  5. Can epigenetic control explain pronounced within plant heterogeneity of meiosis in a translocation trisome of Secale L.?

    NARCIS (Netherlands)

    Sybenga, J.

    2012-01-01

    Meiotic metaphase I configuration frequencies were determined in different tillers of genetically related plants of rye (Secale cereale L.) heterozygous for reciprocal translocation T248W (between chromosome arms 1RS and 6RS) and with an additional (telocentric) arm 1RS. Seventeen different

  6. The chromosomal distributions of Ty1-copia group retrotransposable elements in higher plants and their implications for genome evolution

    Science.gov (United States)

    J.S. (Pat) Heslop-Harrison; Andrea Brandes; Shin Taketa; Thomas Schmidt; Alexander V. Vershinin; Elena G. Alkhimova; Anette Kamm; Robert L. Doudrick; . [and others

    1997-01-01

    Retrotransposons make up a major fraction - sometimes more than 40% - of all plant genomes investigated so far. We have isolated the reverse transcriptase domains of theTyl-copia group elements from several species, ranging in genome size from some 100 Mbp to 23,000 Mbp, and determined the distribution patterns of these retrotransposons on metaphase chromosomes and...

  7. Fish on avian lampbrush chromosomes produces higher resolution gene mapping

    NARCIS (Netherlands)

    Galkina, S.A.; Deryusheva, S.; Fillon, V.; Vignal, A.; Crooijmans, R.P.M.A.; Groenen, M.A.M.; Rodionov, A.V.; Gaginskaya, E.

    2006-01-01

    Giant lampbrush chromosomes, which are characteristic of the diplotene stage of prophase I during avian oogenesis, represent a very promising system for precise physical gene mapping. We applied 35 chicken BAC and 4 PAC clones to both mitotic metaphase chromosomes and meiotic lampbrush chromosomes

  8. National flow cytometry and sorting research resource. Annual progress report, July, 1, 1994--June 30, 1995, Year 12

    Energy Technology Data Exchange (ETDEWEB)

    Jett, J.H.

    1995-04-27

    Research progress utilizing flow cytometry is described. Topics include: rapid kinetics flow cytometry; characterization of size determinations for small DNA fragments; statistical analysis; energy transfer measurements of molecular confirmation in micelles; and enrichment of Mus spretus chromosomes by dual parameter flow sorting and identification of sorted fractions by fluorescence in-situ hybridization onto G-banded mouse metaphase spreads.

  9. Automated dicentric chromosome detection system for radiation dosimetry

    International Nuclear Information System (INIS)

    Xiao Jiang; Luo Yisheng; Cheng Xin; Jiang Li; He Xiaohai

    2000-01-01

    Scoring of dicentric chromosome in human lymphocyte metaphase is the routine method for evaluation of individual dose after radiation accident. Manual microscope scoring methods is tedious and time consuming. The purpose of this subject is to describe a digital image process software of an automated dicentric chromosome detection system the authors developed. Metaphase images are obtained by conventional light microscope equipped with a video camera and converted to digital images. The automatic analysis procedure which developed by C++ computer language including pretreatment of the image, individual chromosome separation and dicentrics scoring. The program was tested using the sample of human blood irradiated with 3.5 Gy γ-rays in vitro. 197 metaphases were analyzed, each of them carries dicentric, about 83% of all dicentrics can be detected correctly, while the value is about 97% for well spread metaphases. An automated dicentric chromosome detection system can give a reliable dose estimation if 80% of all dicentrics are detected correctly. The results show that authors' system can reach this level. As many factors influence the accuracy, such as the quality of cell slide, image capture and pretreatment, much work has to be done before it can be put into practice

  10. Karyotypic differences and evolutionary tendencies of some species ...

    Indian Academy of Sciences (India)

    end concentration 0.20 µg/ml). The metaphase plates were prepared according to the method of Moorhead et al. (1960) adapted for plant cells from in vitro culture. Briefly, cells were centrifuged at 400 g for 5min, treated with 75mM KCl solution ...

  11. A Critical Review of Concepts and Methods Used in Classical Genome Analysis

    DEFF Research Database (Denmark)

    Seberg, Ole; Petersen, Gitte

    1998-01-01

    A short account of the development of classical genome analysis, the analysis of chromosome behaviour in metaphase I of meiosis, primarily in interspecific hybrids, is given. The application of the concept of homology to describe chromosome pairing between the respective chromosomes of a pair dur...

  12. Presenting Mitosis

    Science.gov (United States)

    Roche, Stephanie; Sterling, Donna R.

    2005-01-01

    When the topic of cell division is introduced in the classroom, students can showcase their interpretations of the stages of mitosis by creating a slide show illustrating prophase, metaphase, anaphase, and telophase (see samples in Figure 1). With the help of a computer, they can create a model of mitosis that will help them distinguish the…

  13. Comparison of the aneugenic properties of nocodazole, paclitaxel and griseofulvin in vitro. Centrosome defects and alterations in protein expression profiles.

    Science.gov (United States)

    Zacharaki, Polyxeni; Stephanou, Georgia; Demopoulos, Nikos A

    2013-09-01

    We have comparatively investigated the aneugenic activity of two anticancer drugs, nocodazole (NOC) and paclitaxel (PTX), and the antifungal griseofulvin with promising role in cancer treatment (GF), which affect microtubule dynamics in different ways. The comparison was achieved in HFFF2 human fibroblasts, MCF-7 human breast cancer cells and C2C12 mouse myoblasts, and focused on three issues: (i) induction of chromosome delay by estimation of MN frequency using CREST analysis; (ii) disturbance of spindle organization with Aurora-A/β-tubulin immunofluorescence; and (iii) alterations in the expression of Aurora-A, β- and γ-tubulin by western blotting. They induced chromosome delay, provoked metaphase arrest and promoted microtubule disorganization, reflecting their common characteristic of generating aneuploidy. In particular, NOC induced mainly monopolar metaphases, although PTX induced only multipolar metaphases. GF generated different types of abnormal metaphases, exhibiting cell specificity. Additionally, NOC decreased the expression of Aurora-A and β-tubulin, while the opposite held true for PTX and GF. γ-Tubulin expression was not modulated owing to NOC treatment, whereas PTX and GF increased γ-tubulin expression. Our findings throw a light on the manifestation of the aneugenicity of the studied compounds through centrosome proliferation/separation and protein expression, reflecting their different effects on microtubule dynamics. Copyright © 2012 John Wiley & Sons, Ltd.

  14. Differential dynamics of splicing factor SC35 during the cell cycle

    Indian Academy of Sciences (India)

    We analysed the dynamics of the splicing factor SC35 in interphase and mitotic cells. In HeLa cells expressing green fluorescent protein (GFP)-SC35, this was localized in speckles during interphase and dispersed in metaphase. In telophase, GFP-SC35 was highly enriched within telophase nuclei and also detected in ...

  15. Karyotype analysis of three Solanum plants using combined PI-DAPI ...

    African Journals Online (AJOL)

    ajl yemi

    2011-12-19

    Dec 19, 2011 ... In this study, mitotic metaphase chromosomes of Solanum surattense Burm., Solanum lyratum Thunb. and Solanum photeinocarpum Nakam. were well prepared using an advanced chromosome preparation method. The chromosomes were distinguished by combined PI-DAPI (CPD) staining and double.

  16. Chromosomal distribution of a new centromeric Ty3-gypsy ...

    Indian Academy of Sciences (India)

    nomically important traits, including some that do not exist .... For FISH analysis, probes were labelled with digoxigenin-11-dUTP, and signal detection was performed with fluorescein-conjugated antidigoxigenin antibody according .... FISH using pDbC2 as probe on mitotic metaphase chromosomes of (A) D. breviaristatum.

  17. Cytological effects of oxytocic agents on mitotic chromosomes of ...

    African Journals Online (AJOL)

    These include disturbed prophase, spindle inhibition, star metaphase, chromatid bridge, precocious chromosome and even nuclear disintegration. The percentage of abnormal cells increased with increase in concentration of treatment drugs and duration of treatment. The possible reasons for the low mitotic index, ...

  18. On-line sorting of human chromosomes by centromeric index, and identification of sorted populations by GTG-banding and fluorescent in situ hybridization

    NARCIS (Netherlands)

    Boschman, G. A.; Rens, W.; Manders, E.; van Oven, C.; Barendsen, G. W.; Aten, J. A.

    1990-01-01

    Using slit-scan flow cytometry, the shape of human metaphase chromosomes, as expressed in their centromeric index (CI), and the DNA content of the chromosomes have been used as parameters in bivariate flow karyotyping. The resolution of the DNA vs CI flow karyogram of the larger chromosomes up to

  19. ZEBRAFISH CHROMOSOME-BANDING

    NARCIS (Netherlands)

    PIJNACKER, LP; FERWERDA, MA

    1995-01-01

    Banding techniques were carried out on metaphase chromosomes of zebrafish (Danio rerio) embryos. The karyotypes with the longest chromosomes consist of 12 metacentrics, 26 submetacentrics, and 12 subtelocentrics (2n = 50). All centromeres are C-band positive. Eight chromosomes have a pericentric

  20. A rapid, non-sacrificial chromosome preparation technique for ...

    African Journals Online (AJOL)

    A combination of colchicine, prolonged hypotonic treatment, dissociation of cells followed by Giemsa staining makes it possible to achieve good quality metaphase chromosome spreads using small fish without the use of sterile conditions, centrifuges or sacrificing the specimen. In situations such as the induction of triploidy ...

  1. High resolution FISH on super-stretched flow-sorted plant chromosomes.

    NARCIS (Netherlands)

    Valárik, M.; Bartos, J.; Kovarova, P.; Kubalakova, M.; Jong, de J.H.S.G.M.; Dolezel, J.

    2004-01-01

    A novel high-resolution fluorescence in situ hybridisation (FISH) strategy, using super-stretched flow-sorted plant chromosomes as targets, is described. The technique that allows longitudinal extension of chromosomes of more than 100 times their original metaphase size is especially attractive for

  2. Effect of jasplakinolide on the in vitro maturation of bovine oocytes ...

    African Journals Online (AJOL)

    ... affected the oocyte maturation even after subsequent incubation in normal maturation medium; (5) oocytes at metaphase I were much more sensitive to JAS than oocytes at anaphase I stage. In conclusion, JAS affected polar body extrusion, spindle morphology, microfilament organization and chromosome composition in ...

  3. Physical localization of NORs and ITS length variants in old ...

    Indian Academy of Sciences (India)

    No variation at the number of Ag-NORs per metaphase was found among the 51 durum wheat cultivars, but the PCR-RFLP technique carried out with the restriction enzyme HpaII, allowed the detection of ITS length variants among them. The molecular data was used in order to establish the genetic relationships among ...

  4. Nuclear maturation of immature bovine oocytes after vitrification ...

    African Journals Online (AJOL)

    In the second experiment, effectiveness of both vitrification methods was compared for cryopreservation of immature bovine oocytes. After warming, COCs were cultured in vitro for 24 h. The polar body (PB+) and metaphase-II (MII) stage rates differed significantly among treatment groups. Oocytes vitrified using cryotop ...

  5. Micromechanical study of mitotic chromosome structure

    Science.gov (United States)

    Marko, John

    2011-03-01

    Our group has developed micromanipulation techniques for study of the highly compacted mitotic form of chromosome found in eukaryote cells during cell division. Each metaphase chromosome contains two duplicate centimeter-long DNA molecules, folded up by proteins into cylindrical structures several microns in length. Native chromosomes display linear and reversible stretching behavior over a wide range of extensions (up to 5x native length for amphibian chromosomes), described by a Young modulus of about 300 Pa. Studies using DNA-cutting and protein-cutting enzymes have revealed that metaphase chromosomes behave as a network of chromatin fibers held together by protein-based isolated crosslinks. Our results are not consistent with the more classical model of loops of chromatin attached to a protein-based structural organizer or ``scaffold". In short, our experiments indicate that metaphase chromosomes can be considered to be ``gels" of chromatin; the stretching modulus of a whole chromosome is consistent with stretching of the chromatin fibers contained within it. Experiments using topoisomerases suggest that topological constraints may play an appreciable role in confining chromatin in the metaphase chromosome. Finally, recent experiments on human chromosomes will be reviewed, including results of experiments where chromosome-folding proteins are specifically depleted using siRNA methods. Supported by NSF-MCB-1022117, DMR-0715099, PHY-0852130, DMR-0520513, NCI 1U54CA143869-01 (NU-PS-OC), and the American Heart Association.

  6. Journal of Biosciences | Indian Academy of Sciences

    Indian Academy of Sciences (India)

    Fluorescence in situ hybridization (FISH) is a powerful molecular cytogenetic technique which allows rapid detection of aneuploidies on interphase cells and metaphase spreads. The aim of the present study was to evaluate FISH as a tool in prenatal diagnosis of aneuploidies in high risk pregnancies in an Indian set up.

  7. Insect chromosomes preparing methods for genetic researches

    African Journals Online (AJOL)

    STORAGESEVER

    2009-01-05

    Jan 5, 2009 ... Cytogenetics are almost always based on the examination of the fixed mitotic chromosomes during the analyses of metaphase. During this phase of the cycle of cells, the DNA is folded up and chromatin is strongly condensed. The relative position of the centromere is constant, which means that the ratio of.

  8. Cytogenetic analysis of Astylus antis (Perty, 1830 (Coleoptera, Melyridae: Karyotype, heterochromatin and location of ribosomal genes

    Directory of Open Access Journals (Sweden)

    Ernani de Oliveira Mendes-Neto

    2010-01-01

    Full Text Available Cytogenetic analysis of Astylus antis using mitotic and meiotic cells was performed to characterize the haploid and diploid numbers, sex determination system, chromosome morphology, constitutive heterochromatin distribution pattern and chromosomes carrying nucleolus organizer regions (NORs. Analysis of spermatogonial metaphase cells revealed the diploid number 2n = 18, with mostly metacentric chromosomes. Metaphase I cells exhibited 2n = 8II+Xyp and a parachute configuration of the sex chromosomes. Spermatogonial metaphase cells submitted to C-banding showed the presence of small dots of constitutive heterochromatin in the centromeric regions of nearly all the autosomes and on the short arm of the X chromosome (Xp, as well as an additional band on one of the arms of pair 1. Mitotic cells submitted to double staining with base-specific fluorochromes (DAPI-CMA3 revealed no regions rich in A+T or G+C sequences. Analysis of spermatogonial mitotic cells after sequential Giemsa/AgNO3 staining did not reveal any specific mark on the chromosomes. Meiotic metaphase I cells stained with silver nitrate revealed a strong impregnation associated to the sex chromosomes, and in situ hybridization with an 18S rDNA probe showed ribosomal cistrons in an autosomal bivalent.

  9. Asymmetry in histone H3 variants and lysine methylation between paternal and maternal chromatin of the early mouse zygote.

    NARCIS (Netherlands)

    Heijden, G.W. van der; Dieker, J.W.C.; Derijck, A.H.A.; Muller, S.; Berden, J.H.M.; Braat, D.D.M.; Vlag, J. van der; Boer, P. de

    2005-01-01

    In mammalian fertilization, the paternal genome is delivered to the secondary oocyte by sperm with protamine compacted DNA, while the maternal genome is arrested in meiotic metaphase II. Thus, at the beginning of fertilization, the two gametic chromatin sets are strikingly different. We elaborate on

  10. Cytogenice Studies on Bambara groundnut ( Vigna subterranea (L ...

    African Journals Online (AJOL)

    The study reports the result of analysis of chromosomes of Vigna subterranea performed in the mitotic prometaphase and metaphase stages using conventional techniques. The somatic chromosome study was done using the shoot meristem as the root tips persistently showed very low mitotic indices. The result revealed ...

  11. Chromosome number and meiotic behaviour in Brachiaria jubata ...

    Indian Academy of Sciences (India)

    (f) Metaphase II with precocious chromosome migration to the poles. (g) Late anaphase II with laggard. (h). Tetrad with micronuclei in one microspore. (i) Fertile (dark) and sterile pollen grains (400×). The risks associated with extensive Brachiaria monocul- ture in Brazil are obvious. The two most cultivated vari- eties in the ...

  12. Fulltext PDF

    Indian Academy of Sciences (India)

    2012-08-09

    Aug 9, 2012 ... The 12-year-old patient is the only child of healthy, noncon- ... Cytogenetic studies. Karyotyping was performed on G-banded lymphocyte metaphases from the patient and (subsequently) her parents; at least 16 cells were analysed per individual. In addition, .... 2004 A family with a 17p pericentric inversion:.

  13. Journal of Biosciences | Indian Academy of Sciences

    Indian Academy of Sciences (India)

    It was hyperphosphorylated and showed retarded mobility in cells arrested in metaphase. In vitro experiments showed that it was phosphorylated by CK2 resulting in mobility shift. Using a deletion mutant we found that CK2 phosphorylated PTP-S2 in the C-terminal non-catalytic domain. A heparin sensitive kinase from ...

  14. Journal of Biosciences | Indian Academy of Sciences

    Indian Academy of Sciences (India)

    Fluorescence recovery after photobleaching (FRAP) experiments revealed that the mobility of GFP-SC35 was distinct in different mitotic compartments. Interestingly, the mobility of GFP-SC35 was 3-fold higher in the cytoplasm of metaphase cells compared with interphase speckles, the nucleoplasm or MIGs. Treatment of ...

  15. Journal of Genetics | Indian Academy of Sciences

    Indian Academy of Sciences (India)

    The trypsin inhibitor (ATI) isolated from gastrointestinal nematode Ascaris suum was tested in vitro for induction of chromosome aberrations and sister chromatid exchanges (SCE). Genotoxicity assessment of purified ATI was carried out on metaphase plates received from peripheral blood lymphocyte macroculture (48 h test ...

  16. Morphological, yield, cytological and molecular characterization of a ...

    Indian Academy of Sciences (India)

    J. Lima-Brito et al. to intergenomic recombination since they do not involve chi- asma production. However, the presence of a high frequency of univalents at metaphase I (figure 2,a) could induce Robert- sonian translocations after centric breakage and fusion. In- deed, the absence of chromosome pairing between Hordeum.

  17. Karyological investigations on Erigeron canadensis L. and E. annuus (L. Pers. from natural stands in Poland

    Directory of Open Access Journals (Sweden)

    K. Bijok

    2015-01-01

    Full Text Available The number of chromosomes in metaphases root meristems of Erigeron canadensis L. (2n= 18 and E. annuus (L. Pers. (2n=27 was determined. The chromosomes of both species showed a morphological differentiation - they were heterobrachial or isobrachial; E. annuus had an additional unpaired isobrachial chromosome.

  18. A device for extraction, manipulation and stretching of DNA from single human chromosomes

    DEFF Research Database (Denmark)

    Rasmussen, Kristian Hagsted; Marie, Rodolphe; Moresco, Jacob Lange

    2011-01-01

    by time-lapse imaging; pressure-driven flow was then used to shunt the chromosomal DNA package into a nanoslit. A long linear DNA strand (>1.3 Mbp) was seen to stretch out from the DNA package and along the length of the nanoslit. Delivery of DNA in its native metaphase chromosome package as well...

  19. Chromosomal localization of rDNA genes and genomic organization ...

    Indian Academy of Sciences (India)

    Genomic organization analysis of 5S rDNA revealed two different types of 5S rDNA sequences, 5S type I and 5S type II. Moreover, fluorescence in situ hybridization (FISH) with 5S rDNA probes showed six positive fluorescence signals on six chromosomes of all the analysed metaphases from the three tilapia samples.

  20. Ascorbic acid effects on in vitro maturation of mouse oocyte with or ...

    African Journals Online (AJOL)

    STORAGESEVER

    2009-10-19

    Oct 19, 2009 ... These results indicate that exposure ascorbic acid promotes the development of mouse DOs and COCs from germinal vesicle breakdown (GVBD) to metaphase II (MII) and ... by-product of aerobic metabolism (Miwa and Brand,. 2003). ROS have been recognized to pose a constant threat to cells as they ...

  1. Structure-specific endonucleases

    DEFF Research Database (Denmark)

    Minocherhomji, Sheroy; Hickson, Ian D

    2014-01-01

    Fragile sites are conserved loci predisposed to form breaks in metaphase chromosomes. The inherent instability of these loci is associated with chromosomal rearrangements in cancers and is a feature of cells from patients with chromosomal instability syndromes. One class of fragile sites, the com...

  2. Dicentric assay: Inter-laboratory comparison in Indian laboratories for routine and triage applications

    International Nuclear Information System (INIS)

    Bakkiam, D.; Bhavani, M.; Anantha Kumar, A. Arul; Sonwani, Swetha; Venkatachalam, P.; Sivasubramanian, K.; Venkatraman, B.

    2015-01-01

    An Inter-Laboratory Comparison (ILC) study on Dicentric Chromosome Assay (DCA) was carried out between two Indian biodosimetry labs. Human peripheral blood samples exposed to 10 different doses of X-rays up to 5 Gy were shared between the labs to generate calibration data. Validation of calibration curves was done by dose estimation of coded samples exposed to X- or gamma radiation. Reliability of the DCA data for triage application was evaluated by scoring 20, 50 and 100 metaphases in the dose range of 0.5–3.0 Gy. No significant difference was observed between labs regarding the established calibration data as well as the DCA triage dose assessments. Scoring of 20 metaphases (MP) was adequate to detect radiation exposure of >2 Gy whereas 50 MP were sufficient to determine exposures of 0.5 Gy. Both labs performed the DCA in a reliable manner and made the first step in setting up a biodosimetry network in India. - Highlights: • This is the first Inter-Laboratory Comparison (ILC) study from India on Dicentric Chromosomal Assay (DCA) for both routine and triage biodosimetry applications. • No significant difference in the in vitro dose response curve parameters (slope, intercept and curvature) between labs. • Validation of calibration curves was done by blind trial method using samples exposed to different doses of X-rays and gamma rays. • Triage analysis was done by scoring 20, 50 and 100 metaphases for dose determination. • Scoring 20 metaphases is adequate to detect radiation exposure of 2 Gy and 50 metaphases is sufficient to determine radiation exposures of as low as 0.5 Gy. • The feasibility of formation of biodosimetry network in India which is essential especially when the country is aiming at capacity building in nuclear energy is demonstrated

  3. Cytogenic follow-up studies in six radiation accident victims 16 and 17 years post-exposure

    International Nuclear Information System (INIS)

    Littlefield, L.G.; Joiner, E.E.

    1978-01-01

    Detailed cytogenetic evaluations were recently conducted in cultured lymphocytes from six men accidentally exposed to fission neutron and gamma radiation in 1958 (dose range 22.8-365 rads). Two-day lymphocyte cultures stimulated with phytohaemagglutinin (PHA) and seven-day cultures stimulated with pokeweed mitogen (PWM) were initiated on all six men for routine microscopic and karyotypic analysis. PHA-stimulated cultures from the two men with the highest exposures were also evaluated using Giemsa-banding procedures. Approximately 9% of the metaphases in the PHA cultures and 11% of the metaphases in the PWM cultures were found to have residual radiation-induced aberrations (symmetrical and asymmetrical exchanges and deletions). In both the PHA and PWM cultures the highest frequencies of lesions were observed in preparations from three men with the highest radiation exposures. Among the abnormal metaphases from the various PHA cultures five possible clones were identified (two or more cells having apparently identical lesions) whereas no stemlines were detected in the PWM cultures. The frequency of stable lesions in 92 banded metaphases from the two men with the highest exposures did not differ significantly from the frequency of lesions detected in 300 metaphases evaluated with routine staining and karyotypic procedures. These data suggest that radiation-damaged lymphocytes responsive to mitogenic stimulation by PHA and PWM may survive for many years following exposure, that the frequency of lymphocytes with persistent aberrations can be roughly correlated with dose, and that in some instances cells bearing 'stable' radiation-induced lesions may propagate in vivo. The findings also show that banding procedures, compared with routine microscopic and karyotypic methods, do not significantly improve the rate of detection of symmetrical lesions in lymphocytes from irradiated persons

  4. Semiautomatic digital imaging system for cytogenetic analysis

    International Nuclear Information System (INIS)

    Chaubey, R.C.; Chauhan, P.C.; Bannur, S.V.; Kulgod, S.V.; Chadda, V.K.; Nigam, R.K.

    1999-08-01

    The paper describes a digital image processing system, developed indigenously at BARC for size measurement of microscopic biological objects such as cell, nucleus and micronucleus in mouse bone marrow; cytochalasin-B blocked human lymphocytes in-vitro; numerical counting and karyotyping of metaphase chromosomes of human lymphocytes. Errors in karyotyping of chromosomes by the imaging system may creep in due to lack of well-defined position of centromere or extensive bending of chromosomes, which may result due to poor quality of preparation. Good metaphase preparations are mandatory for precise and accurate analysis by the system. Additional new morphological parameters about each chromosome have to be incorporated to improve the accuracy of karyotyping. Though the experienced cytogenetisist is the final judge; however, the system assists him/her to carryout analysis much faster as compared to manual scoring. Further, experimental studies are in progress to validate different software packages developed for various cytogenetic applications. (author)

  5. Cytogenetical effects of gamma rays and maleic hydrazide in mung bean

    International Nuclear Information System (INIS)

    Grover, I.S.; Tejpaul, S.K.

    1982-01-01

    The cytological effects of gamma rays, maleic hydrazide and gamma rays+MH were examined on Vigna radiata (L) Wilczek. The disturbed mitotic behaviour was noticed following gamma irradiation or chemical mutagenic treatment. The sticky chromosomes, fragments and ring chromosomes were noticed at metaphase. The laggards and bridges were noticed at anaphase. The chromosomal aberrartions were found to be significantly correlated with dose. The combined treatment enhanced chromosomal aberrations. Similarly the meiotic process was also affected. The quadrivalents presumably due to translocations were occasionally encountered at metaphase-I. Irregular disjunction of chromosomes at anaphase-I accompanied by laggards was observed. The statistical analysis revealed that aberration is a function of dose. A reduction in pollen fertility as well as seed set was noticed. The meiotic analysis of the plants of M 2 generation revealed the presence of chromosomal abnormalities. (author)

  6. Cytotaxonomy study of four populations of Astragalus anserinifolius Boiss. of section Malacothrix Bunge from Iran

    Directory of Open Access Journals (Sweden)

    Massoud Ranjbar

    2013-01-01

    Full Text Available In this research, meiotic chromosome number and the behavior of four populations of Astragalus anserinifolius Boiss. of Astragalus sect. Malacothrix were studied. All wild populations were diploid and showed 2n=2x=16 chromosome number, consistent with the proposed base number of x=8 from IPCN. Although all taxa displayed regular bivalent pairing and chromosome segregation at meiosis, some meiotic abnormalities included varied degrees of fragmented and sticky chromosomes in metaphase I, polynucleate and a variable number of laggards, forwarded chromosomes and bridges in anaphase I/telophase I, asynchronous nucleus and precocious chromosome migration in metaphase II and laggards, bridges and cytomixis in anaphase II/telophase II were observed.

  7. Gonadotropin-releasing hormone agonist trigger in oocyte donors co-treated with a gonadotropin-releasing hormone antagonist

    DEFF Research Database (Denmark)

    Vuong, T. N. L.; Ho, M. T.; Ha, T. D.

    2016-01-01

    -35 years, body mass index [BMI] hormone level >1.25 ng/mL, and antral follicle count >= 6). Intervention(s): Ovulation trigger with 0.2, 0.3, or 0.4 mg triptorelin in a GnRH antagonist cycle. Main Outcome Measure(s): The primary end point was number of metaphase II oocytes...... to number of metaphase II oocytes (16.0 +/- 8.5, 15.9 +/- 7.8, and 14.7 +/- 8.4, respectively), embryos (13.2 +/- 7.8, 11.7 +/- 6.9, 11.8 +/- 7.0), and number of top-quality embryos (3.8 +/- 2.9, 3.6 +/- 3.0, 4.1 +/- 3.0). Luteinizing hormone levels at 24 hours and 36 hours after trigger was significantly...

  8. Inhibition of clathrin by pitstop 2 activates the spindle assembly checkpoint and induces cell death in dividing HeLa cancer cells

    Directory of Open Access Journals (Sweden)

    Smith Charlotte M

    2013-01-01

    Full Text Available Abstract Background During metaphase clathrin stabilises the mitotic spindle kinetochore(K-fibres. Many anti-mitotic compounds target microtubule dynamics. Pitstop 2™ is the first small molecule inhibitor of clathrin terminal domain and inhibits clathrin-mediated endocytosis. We investigated its effects on a second function for clathrin in mitosis. Results Pitstop 2 did not impair clathrin recruitment to the spindle but disrupted its function once stationed there. Pitstop 2 trapped HeLa cells in metaphase through loss of mitotic spindle integrity and activation of the spindle assembly checkpoint, phenocopying clathrin depletion and aurora A kinase inhibition. Conclusions Pitstop 2 is therefore a new tool for investigating clathrin spindle dynamics. Pitstop 2 reduced viability in dividing HeLa cells, without affecting dividing non-cancerous NIH3T3 cells, suggesting that clathrin is a possible novel anti-mitotic drug target.

  9. Homogeneously staining regions (HSR) in a human malignant melanoma.

    Science.gov (United States)

    Brieux de Salum, S; Slavutsky, I; Besuschio, S; Pavlovsky, A A

    1984-01-01

    A case of nodular malignant melanoma (level V of Clark's classification) with homogeneously staining regions (HSR) on the long arm of one chromosome #2 is described. Ultrastructural observation of melanosomic and promelanosomic granules near Golgi's vesicles confirmed the histologic diagnosis. Chromosome analysis was performed on nine metaphases from a bone marrow sample and 76 metaphases from culture of the malignant skin tumor. G-banding revealed the presence of a clone with trisomy #8 and another cell line with the HSR marker. This is the first report of HSR in human melanoma cells. As HSR has been found only in malignant cells, we believe that among the many factors that influence the patients' clinical evolution and poor response to treatment, the genic imbalance is of the utmost importance.

  10. Contamination of persons occupationally exposed to natural radioactivity in a coal fired power plant

    International Nuclear Information System (INIS)

    Bauman, A.; Horvat, D.

    1980-01-01

    Contamination of occupationally exposed subjects with natural radioactivity in a coal fired power plant at levels of 500 mrem/year was detected. The level of 210 Pb in urine varied from 2.29-14.47 pCi/l. These values were arrived at after subtracting a blank value of 1.05 pCi 210 Pb obtained from a control group. Structural chromosomal aberrations, completely missing in the control group, were detected in the exposed subjects. Approximately 6-10% of the metaphases of occupationally exposed subjects were found to have aberrations which were probably radiation induced. These included symmetrical and asymmetrical exchanges and numerical aberrations. In the control aroup aberrations were found in 1.4-4% of the metaphases, but these were only deletions. (H.K.)

  11. Hidden chromosomal abnormalities in pleuropulmonary blastomas identified by multiplex FISH

    International Nuclear Information System (INIS)

    Quilichini, Benoit; Andre, Nicolas; Bouvier, Corinne; Chrestian, Marie-Anne; Rome, Angelique; Intagliata, Dominique; Coze, Carole; Lena, Gabriel; Zattara, Helene

    2006-01-01

    Pleuropulmonary blastoma (PPB) is a rare childhood dysontogenetic intrathoracic neoplasm associated with an unfavourable clinical behaviour. We report pathological and cytogenetic findings in two cases of PPB at initial diagnosis and recurrence. Both tumors were classified as type III pneumoblastoma and histological findings were similar at diagnosis and relapse. In both cases, conventional cytogenetic techniques revealed complex numerical and structural chromosomal abnormalities. Molecular cytogenetic analysis (interphase/metaphase FISH and multicolor FISH) identified accurately chromosomal aberrations. In one case, TP53 gene deletion was detected on metaphase FISH. To date, only few cytogenetic data have been published about PPB. The PPB genetic profile remains to be established and compared to others embryonal neoplasia. Our cytogenetic data are discussed reviewing cytogenetics PPBs published cases, illustrating the contribution of multicolor FISH in order to identify pathogenetically important recurrent aberrations in PPB

  12. Protocol for chromosome-specific probe construction using PRINS, micromanipulation and DOP-PCR techniques

    Directory of Open Access Journals (Sweden)

    PAULO Z. PASSAMANI

    2017-12-01

    Full Text Available ABSTRACT Chromosome-specific probes have been widely used in molecular cytogenetics, being obtained with different methods. In this study, a reproducible protocol for construction of chromosome-specific probes is proposed which associates in situ amplification (PRINS, micromanipulation and degenerate oligonucleotide-primed PCR (DOP-PCR. Human lymphocyte cultures were used to obtain metaphases from male and female individuals. The chromosomes were amplified via PRINS, and subcentromeric fragments of the X chromosome were microdissected using microneedles coupled to a phase contrast microscope. The fragments were amplified by DOP-PCR and labeled with tetramethyl-rhodamine-5-dUTP. The probes were used in fluorescent in situ hybridization (FISH procedure to highlight these specific regions in the metaphases. The results show one fluorescent red spot in male and two in female X chromosomes and interphase nuclei.

  13. Protocol for chromosome-specific probe construction using PRINS, micromanipulation and DOP-PCR techniques.

    Science.gov (United States)

    Passamani, Paulo Z; Carvalho, Carlos R; Soares, Fernanda A F

    2018-01-01

    Chromosome-specific probes have been widely used in molecular cytogenetics, being obtained with different methods. In this study, a reproducible protocol for construction of chromosome-specific probes is proposed which associates in situ amplification (PRINS), micromanipulation and degenerate oligonucleotide-primed PCR (DOP-PCR). Human lymphocyte cultures were used to obtain metaphases from male and female individuals. The chromosomes were amplified via PRINS, and subcentromeric fragments of the X chromosome were microdissected using microneedles coupled to a phase contrast microscope. The fragments were amplified by DOP-PCR and labeled with tetramethyl-rhodamine-5-dUTP. The probes were used in fluorescent in situ hybridization (FISH) procedure to highlight these specific regions in the metaphases. The results show one fluorescent red spot in male and two in female X chromosomes and interphase nuclei.

  14. PTEN regulates EG5 to control spindle architecture and chromosome congression during mitosis

    Science.gov (United States)

    He, Jinxue; Zhang, Zhong; Ouyang, Meng; Yang, Fan; Hao, Hongbo; Lamb, Kristy L.; Yang, Jingyi; Yin, Yuxin; Shen, Wen H.

    2016-01-01

    Architectural integrity of the mitotic spindle is required for efficient chromosome congression and accurate chromosome segregation to ensure mitotic fidelity. Tumour suppressor PTEN has multiple functions in maintaining genome stability. Here we report an essential role of PTEN in mitosis through regulation of the mitotic kinesin motor EG5 for proper spindle architecture and chromosome congression. PTEN depletion results in chromosome misalignment in metaphase, often leading to catastrophic mitotic failure. In addition, metaphase cells lacking PTEN exhibit defects of spindle geometry, manifested prominently by shorter spindles. PTEN is associated and co-localized with EG5 during mitosis. PTEN deficiency induces aberrant EG5 phosphorylation and abrogates EG5 recruitment to the mitotic spindle apparatus, leading to spindle disorganization. These data demonstrate the functional interplay between PTEN and EG5 in controlling mitotic spindle structure and chromosome behaviour during mitosis. We propose that PTEN functions to equilibrate mitotic phosphorylation for proper spindle formation and faithful genomic transmission. PMID:27492783

  15. Replacement of radiography with ultrasonic phased array for feeder tubes in CANDU reactors using ASME code case N-659-2

    International Nuclear Information System (INIS)

    Simmons, R.; Bower, Q.; Arseneau, S.

    2013-01-01

    In this paper we will discuss phased array technology for the replacement of radiography on new construction projects in the nuclear industry. Specifically, through the implementation of A.S.M.E. code N-659-2 and MetaPhase phased array services. Phased Array is not considered a new technique on in service welds in the nuclear industry; however it was unprecedented on new construction welds and required significant investment in regulatory approval (C.N.S.C.), technology research and development, regulatory, client and technician training for successful service implementation. This paper will illustrate the abilities and limitations associated in replacing radiography with MetaPhase, as well as the substantial benefits relative to increased production, improved weld quality, enhanced safety and overall project cost savings. (author)

  16. First report of a patient with a mixoploidy 47,XXX/94,XXXXXX.

    Science.gov (United States)

    Rodríguez Criado, G; Galán Gómez, E; Tizzano, E F; García Rodríguez, E; Gómez de Terreros, I

    2007-01-01

    We present a 16 years old female with a chromosomal mixoploidy and multiple phenotypic anomalies. Peripheral blood G-band karyotype was 47,XXX and her skin fibroblast karyotype revealed a mosaic with a 47,XXX cell line in 88% of metaphases and a 94,XXXXXX cell line in 12% of metaphases, consistent with a hypertetraploidy. The most prominent clinical signs were: short stature, left upper limb asymmetry, senile-like appearance, generalized hypertrichosis, and small hands and feet. Radiological examination showed bone dysplasia. The result of molecular studies demonstrated that the patient inherited the two X chromosomes from the mother and one from the father, indicating that her 47,XXX trisomy resulted from an oogenesis error in the first meiotic division. The 94,XXXXXX cell line was likely the result of a cytokinesis error. To our knowledge, this is the first documented patient with a trisomy and a hypertetraploidy.

  17. Transcriptional intermediary factor 1γ binds to the anaphase-promoting complex/cyclosome and promotes mitosis

    DEFF Research Database (Denmark)

    Sedgwick, G.G.; Townsend, K.; Martin, A.

    2013-01-01

    The anaphase-promoting complex/cyclosome (APC/C) is an ubiquitin ligase that functions during mitosis. Here we identify the transcriptional regulator, transcriptional intermediary factor 1γ, TIF1γ, as an APC/C-interacting protein that regulates APC/C function. TIF1γ is not a substrate for APC....../C-dependent ubiquitylation but instead, associates specifically with the APC/C holoenzyme and Cdc20 to affect APC/C activity and progression through mitosis. RNA interference studies indicate that TIF1γ knockdown results in a specific reduction in APC/C ubiquitin ligase activity, the stabilization of APC/C substrates......, and an increase in the time taken for cells to progress through mitosis from nuclear envelope breakdown to anaphase. TIF1γ knockdown cells are also characterized by the inappropriate presence of cyclin A at metaphase, and an increase in the number of cells that fail to undergo metaphase-to-anaphase transition...

  18. Unmet needs in automated cytogenetics

    International Nuclear Information System (INIS)

    Bender, M.A.

    1976-01-01

    Though some, at least, of the goals of automation systems for analysis of clinical cytogenetic material seem either at hand, like automatic metaphase finding, or at least likely to be met in the near future, like operator-assisted semi-automatic analysis of banded metaphase spreads, important areas of cytogenetic analsis, most importantly the determination of chromosomal aberration frequencies in populations of cells or in samples of cells from people exposed to environmental mutagens, await practical methods of automation. Important as are the clinical diagnostic applications, it is apparent that increasing concern over the clastogenic effects of the multitude of potentially clastogenic chemical and physical agents to which human populations are being increasingly exposed, and the resulting emergence of extensive cytogenetic testing protocols, makes the development of automation not only economically feasible but almost mandatory. The nature of the problems involved, and acutal of possible approaches to their solution, are discussed

  19. Radiation-induced chromosome aberrations in bone marrow cells leading to acute myeloid leukemia in mouse

    International Nuclear Information System (INIS)

    Nobuhiko Ban; Tomoko Kusama

    1996-01-01

    It is well known that radiation-induced acute myeloid leukemia (RI-AML) in mice is charaterized by deletion and/or rearrangement of chromosome 2. While chromosome 2 has been suspected to be a target of RI-AML, radiation-sensitive site of the chromosome might be implicated in the leukemogenesis. There were few cytogenetical studies, however, focusing on chromosomal rearrangements shortly after irradiation, and little was known about the frequency and pattern of chromosome 2 aberrations during the early period. In this study, metaphase samples were prepared from whole-body irradiated mice 24 hours after irradiation, most of the cells considered to be in the first mitotic stage. Distribution of chromosomal breakpoints on the metaphase samples were analyzed to study the relationship between chromosome aberrations and RI-AML. (author)

  20. The effect of X-irradiation on the fertility and the induction of meiotic chromosome rearrangements in mice and their first generation

    International Nuclear Information System (INIS)

    Savkovic, N.; Pecevski, J.; Maric, N.; Radivojevic, D.

    1978-01-01

    The effect of whole-body and local irradiation with a dose of 600 X-rays on the induction of chromosomal translocations in Diakinesis-Metaphase I of meiosis in treated and F 1 males and their fertility have been examined. Our results showed the high percentage of mortality in whole-body irradiated mice. The percentage of fertility was 25% in whole-body, and 93,7% in locally irradiated males. The testis weight was also reduced. The percentage of chromosomal translocations in Diakinesis-Metaphase I of meiosis was greater after whole-body than after local irradiation. In F 1 males both types of irradiation induced chromosomal translocations. (orig.) [de

  1. Experimental study of mutagenous and mitosis modifying activity of silver nanoparticles

    Directory of Open Access Journals (Sweden)

    B. S. Kirbik

    2015-01-01

    Full Text Available Mutagenous and mitosis modifying impact of silver nanoparticles has been studied on outbred mice. Nanoparticles were of round shape with dimensions of 5-50 nm, size of generated organic shell of 2-5 nm, the quantity in 1 mcm3 makes 120-270. Metaphasic analysis of mice bone marrow cells was used as a testing technique. The frequency of chromosome aberrations and mitotic index of preparations were accounted. During single intraperitoneal administration of the agent in the dose of 250 mcg/kg the silver nanoparticles demonstrated mitosis stimulating activity. No mutagenous effect of silver nanoparticles by daily administration for 4 days of 25 mcg/kg and single administration in the dose of 250 mcg/kg has been registered, but there is statistically insignificant tendency of aberrant metaphases increase. Consequently silver nanoparticles in the investigated doses demonstrated no mutagenous activity and can be considered safe for mammalian cells.

  2. Molecular Cytogenetic Analysis of Spontaneous Interspecific Hybrid Between Oryza sativa and Oryza minuta

    Directory of Open Access Journals (Sweden)

    Chuan-deng YI

    2008-12-01

    Full Text Available Genomic in situ hybridization (GISH is a powerful tool to characterize parental chromosomes in interspecific hybrids, including the behaviour of autosynapsis and chromosome pairing. It was used to distinguish the chromosomes of Oryza sativa from wild species in a spontaneous interspecific hybrid and to investigate the chromosome pairing at metaphase I in meiosis of the hybrid in this study. The hybrid was a triploid with 36 chromosomes according to the chromosome number investigated in mitosis of root tips. During metaphase I of meiosis in the hybrid, less chromosome pairing was observed and most of the chromosomes existed as univalent. Based on GISH and FISH (Fluorescent in situ hybridization analyses, the chromosomes of the hybrid were composed of genomes A, B and C. Thus, it was believed that the hybrid was the result of natural hybridization between cultivated rice and wild species O. minuta which was planted in experimental fields.

  3. Prenatal diagnosis and molecular cytogenetic characterization of a de novo 4.858-Mb microdeletion in 15q14 associated with ACTC1 and MEIS2 haploinsufficiency and tetralogy of Fallot

    Directory of Open Access Journals (Sweden)

    Chih-Ping Chen

    2016-04-01

    Conclusion: Fetuses with 15q14 microdeletion may present TOF on the second-trimester ultrasound. aCGH and metaphase FISH are useful for rapid prenatal diagnosis of 15q14 microdeletion associated with TOF. A prenatal diagnosis of TOF should include a differential diagnosis of 15q14 microdeletion in addition to 22q11.2 deletion syndrome and other microdeletion syndromes.

  4. Balanced Autosomal Translocations in Two Women Reporting Recurrent Miscarriage

    OpenAIRE

    Arumugam, Brindha; Samuel, Chandra R; Thyagarajan, Santhiya Sathiyavedu

    2016-01-01

    Spontaneous abortion or loss of fetus prior to 20 weeks of gestation is observed in 15-20% of clinically recognized pregnancies. Recurrent Miscarriage (RM) is defined as three or more consecutive pregnancy losses and it affects 1-2% of women. Parental chromosomal rearrangements account for 2-5% of RM. This report describes two couples with a clinical history of RM who were subjected to conventional cytogenetic analysis to ascertain the chromosomal aetiology. Analysis of GTG-banded metaphases ...

  5. Abnormal Nuclear Shape in Solid Tumors Reflects Mitotic Instability

    OpenAIRE

    Gisselsson, David; Björk, Jonas; Höglund, Mattias; Mertens, Fredrik; Dal Cin, Paola; Åkerman, Måns; Mandahl, Nils

    2001-01-01

    Abnormalities in nuclear morphology are frequently observed in malignant tissues but the mechanisms behind these phenomena are still poorly understood. In this study, the relation between abnormal nuclear shape and chromosomal instability was explored in short-term tumor cell cultures. Mitotically unstable ring and dicentric chromosomes were identified by fluorescence in situ hybridization at metaphase and subsequently localized in interphase nuclei from five malignant soft tissue tumors. The...

  6. Cytogenetic characterization of Nigerian indigenous pig

    African Journals Online (AJOL)

    STORAGESEVER

    2009-09-15

    Sep 15, 2009 ... 1 h before harvesting, colchicine 0.8 µg/ml was added to arrest the cells at metaphase stage. After centrifu- gation at 1000 rpm for 10 min, the blood lymphocyte layer was collected and transferred into clean dry test-tubes. 5-10 ml of 0.075% of hypotonic solution was added, re- centrifuged and fixed three ...

  7. Improved cytometry - Phase II

    International Nuclear Information System (INIS)

    McLean, J.R.N.

    1995-09-01

    One shortcoming of biological dosimetry based on the detection and quantitation of chromosome aberrations, is the labor intensive nature of the analysis. State-of-the-art imaging software cannot recognize the wide variety of shapes that can be assumed by dicentric chromosomes nor the algorithms available to accommodate translocation analysis. In this project, relatively simple computer routines have been used to reduce the time necessary to perform the repetitive tasks associated with radiation dosimetry. Repetitive tasks that can be computerized include data reduction and collation, verification of the number of chromosomes in each metaphase spread and, with the help of a motorized microscope stage and computer-assisted focus control, the automated location of metaphase chromosome spreads. The task of automated metaphase location is addressed in this project. The tasks of data reduction and computer-assisted chromosome counting have been addressed in a previous report to the Atomic Energy Control Board (AECB) research project no.7.164.1. The preliminary evaluation of the performance of the metaphase finder, indicates a false positive rate of less than or equal to 7% and a false negative rate of less than or equal to 23%, using a limited data set of 1289 events and threshold values at 25, 26, 27, or 28. Over the 4 threshold levels, the average system sensitivity was calculated to be 75% and positive predictability to be 85%. Sensitivity is the fraction of real events which are correctly detected, and positive predictability, the fraction of detections which are real events. In a perfect detector, these values would be 100%. (author). 5 refs., 1 tab

  8. Ultraviolet light induced changes in the organization of mitotic chromosomes in porphyria cutanea tarda

    International Nuclear Information System (INIS)

    Rasko, I.; Went, M.; Johnson, R.T.; University Medical School, Szeged; Cambridge Univ.

    1977-01-01

    Chromosome decondensation or attenuation can be induced with ultraviolet light, particularly in the presence of inhibitors of semiconservative DNA synthesis, such as hydroxyurea. The rate of decondensation of metaphase chromosomes from lymphocyte cultures of healthy controls and patients with porphyria cutanea tarda has been compared after ultraviolet irradiation in the presence of hydroxyurea. Chromosomes of the patients attenuated more readily than the control ones at all UV doses. The possibilities causing such differences in the profile of chromosome decondensation are discussed

  9. Multiplication of human NHIK 3025 cells exposed to porphyrins in combination with light.

    OpenAIRE

    Christensen, T.

    1981-01-01

    Cells from the established line NHIK 3025 were exposed to haematoporphyrin derivative and light. After this photodynamic treatment the first interphase of surviving cells was prolonged. Furthermore, a pronounced effect on the progression through the first mitosis was observed. Mainly the duration of metaphase was increased. Some of the cells were irreversibly arrested in mitosis and the cells that were able to complete mitosis after treatment multiplied in the subsequent generations at the sa...

  10. Time-sequential observation of spindle and phragmoplast orientation in BY-2 cells with altered cortical actin microfilament patterning

    OpenAIRE

    Kojo, Kei H; Yasuhara, Hiroki; Hasezawa, Seiichiro

    2014-01-01

    Precise division plane determination is essential for plant development. At metaphase, a dense actin microfilament meshwork appears on both sides of the cell center, forming a characteristic cortical actin microfilament twin peak pattern in BY-2 cells. We previously reported a strong correlation between altered cortical actin microfilament patterning and an oblique mitotic spindle orientation, implying that these actin microfilament twin peaks play a role in the regulation of mitotic spindle ...

  11. A 62-kD protein required for mitotic progression is associated with the mitotic apparatus during M-phase and with the nucleus during interphase

    OpenAIRE

    Johnston, Jennifer A.; Sloboda, Roger D.

    1992-01-01

    A protein of 62 kD is a substrate of a calcium/calmodulin-dependent protein kinase, and both proteins copurify with isolated mitotic apparatuses (Dinsmore, J. H., and R. D. Sloboda. 1988. Cell. 53:769- 780). Phosphorylation of the 62-kD protein increases after fertilization; maximum incorporation of phosphate occurs during late metaphase and anaphase and correlates directly with microtubule disassembly as determined by in vitro experiments with isolated mitotic apparatuses. Because 62-kD prot...

  12. The impact of trisomy 12, retinoblastoma gene and P53 in prognosis of B-cell chronic lymphocytic leukemia.

    Science.gov (United States)

    AbdelSalam, M; El Sissy, A; Samra, M A; Ibrahim, S; El Markaby, D; Gadallah, F

    2008-06-01

    Routine cytogenetic analysis frequently fails to identify an abnormal clone in B-cell lymphocytic leukaemia (B-CLL) due to poor response to mitogen stimulation. Fluorescence in situ hybridization (FISH) suggest that chromosomal abnormalities occur more frequently, most commonly trisomy 12, retinoblastoma gene deletion (Rb1 gene) and P53 gene deletion. 30 patients with B-CLL were enrolled in the trial from two centers in Cairo, Egypt during the period May 2000 to January 2002. Karyotyping and FISH assessment for possible chromosomal abnormalities (trisomy 12, Rb1 gene and P53 gene) were done at initial diagnosis. Results of cytogenetic abnormalities were correlated with clinical picture and survival. The median age was 57.4 years (range 40-75). Karyotyping technique showed that no metaphase could be detected in 30%, metaphase with normal karyotyping was observed in 63% and cytogenetic abnormalities were detected in two cases (one trisomy 12 and one deletion in chromosome 13). FISH examination of interphase and metaphase nuclei revealed cytogenetic abnormalities in 15 cases (50%), trisomy 12 in 9 (30%), Rb1 gene deletion in 5 (17%) and P53 gene deletion in 3. At diagnosis, patients with trisomy 12 were significantly associated with advanced stage and absolute lymphocyte count of >or=30,000/mm(3). Univariate analysis showed that absolute lymphocyte count >or=30,000/mm(3) (p=0.004) and trisomy 12 (p=0.024) were associated with poor progression free survival. Interphase and metaphase FISH studies improve the cytogenetic diagnosis of chromosomal abnormalities in B-CLL. Lymphocytosis and trisomy 12 may be a good indicator of poor prognosis.

  13. [Partial trisomy of chromosome 15 with new phenotypic manifestations].

    Science.gov (United States)

    Mar González, J; Llaurado Robles, R A; Cabrera Rivas, T; Lantigua Cruz, A; Rodríguez Verdecia, B

    1994-01-01

    A patient with a 15 partial trisomy and a 4 target chromosome in 100% of metaphases is presented. Phenotypic manifestations not previously described were observed such as macrocephally, long face, low implantation of ears, narrow forehead, epicanthal fold, copious eyebrows and synophrys, short nasolabial distance, convergent strabismus, delayed bucal eruption, long neck, hypertrophy of thenar and hypothenar bulging and articular hypermobility. The eyeground was degeneratively myopic. This case makes more extensive the variety of clinical manifestations of this disease.

  14. When the genome plays dice: circumvention of the spindle assembly checkpoint and near-random chromosome segregation in multipolar cancer cell mitoses.

    Directory of Open Access Journals (Sweden)

    David Gisselsson

    Full Text Available BACKGROUND: Normal cell division is coordinated by a bipolar mitotic spindle, ensuring symmetrical segregation of chromosomes. Cancer cells, however, occasionally divide into three or more directions. Such multipolar mitoses have been proposed to generate genetic diversity and thereby contribute to clonal evolution. However, this notion has been little validated experimentally. PRINCIPAL FINDINGS: Chromosome segregation and DNA content in daughter cells from multipolar mitoses were assessed by multiphoton cross sectioning and fluorescence in situ hybridization in cancer cells and non-neoplastic transformed cells. The DNA distribution resulting from multipolar cell division was found to be highly variable, with frequent nullisomies in the daughter cells. Time-lapse imaging of H2B/GFP-labelled multipolar mitoses revealed that the time from the initiation of metaphase to the beginning of anaphase was prolonged and that the metaphase plates often switched polarity several times before metaphase-anaphase transition. The multipolar metaphase-anaphase transition was accompanied by a normal reduction of cellular cyclin B levels, but typically occurred before completion of the normal separase activity cycle. Centromeric AURKB and MAD2 foci were observed frequently to remain on the centromeres of multipolar ana-telophase chromosomes, indicating that multipolar mitoses were able to circumvent the spindle assembly checkpoint with some sister chromatids remaining unseparated after anaphase. Accordingly, scoring the distribution of individual chromosomes in multipolar daughter nuclei revealed a high frequency of nondisjunction events, resulting in a near-binomial allotment of sister chromatids to the daughter cells. CONCLUSION: The capability of multipolar mitoses to circumvent the spindle assembly checkpoint system typically results in a near-random distribution of chromosomes to daughter cells. Spindle multipolarity could thus be a highly efficient

  15. When the genome plays dice: circumvention of the spindle assembly checkpoint and near-random chromosome segregation in multipolar cancer cell mitoses.

    Science.gov (United States)

    Gisselsson, David; Håkanson, Ulf; Stoller, Patrick; Marti, Dominik; Jin, Yuesheng; Rosengren, Anders H; Stewénius, Ylva; Kahl, Fredrik; Panagopoulos, Ioannis

    2008-04-02

    Normal cell division is coordinated by a bipolar mitotic spindle, ensuring symmetrical segregation of chromosomes. Cancer cells, however, occasionally divide into three or more directions. Such multipolar mitoses have been proposed to generate genetic diversity and thereby contribute to clonal evolution. However, this notion has been little validated experimentally. Chromosome segregation and DNA content in daughter cells from multipolar mitoses were assessed by multiphoton cross sectioning and fluorescence in situ hybridization in cancer cells and non-neoplastic transformed cells. The DNA distribution resulting from multipolar cell division was found to be highly variable, with frequent nullisomies in the daughter cells. Time-lapse imaging of H2B/GFP-labelled multipolar mitoses revealed that the time from the initiation of metaphase to the beginning of anaphase was prolonged and that the metaphase plates often switched polarity several times before metaphase-anaphase transition. The multipolar metaphase-anaphase transition was accompanied by a normal reduction of cellular cyclin B levels, but typically occurred before completion of the normal separase activity cycle. Centromeric AURKB and MAD2 foci were observed frequently to remain on the centromeres of multipolar ana-telophase chromosomes, indicating that multipolar mitoses were able to circumvent the spindle assembly checkpoint with some sister chromatids remaining unseparated after anaphase. Accordingly, scoring the distribution of individual chromosomes in multipolar daughter nuclei revealed a high frequency of nondisjunction events, resulting in a near-binomial allotment of sister chromatids to the daughter cells. The capability of multipolar mitoses to circumvent the spindle assembly checkpoint system typically results in a near-random distribution of chromosomes to daughter cells. Spindle multipolarity could thus be a highly efficient generator of genetically diverse minority clones in transformed cell

  16. Biological effects of 137Cs, incorporated into organism of rats

    International Nuclear Information System (INIS)

    Monakhov, A.S.; Strekalov, S.A.; Sokolov, A.V.; Aver'yanova, T.K.

    1987-01-01

    Results of investigating mutagenous and hemotoxic effects of 137 Cs on blood lymphocytes of rats are presented. 137 Cs was orally administrated into organism of rats as 270 kBq/g chloride solution. 137 Cs mutagenous effect was studied on metaphase plates of rat blood lymphocytes in course of rats lifetime experiment. It is stated that 137 Cs inducing severe disturbances of genetic material in a great quantity of blood lymphocytes, causes their total killing

  17. Field-flow fractionation of chromosomes

    Energy Technology Data Exchange (ETDEWEB)

    Giddings, J.C.

    1990-09-01

    Research continued on field flow fractionation of chromosomes. Progress in the past year can be organized into three main categories: (1) chromosome sample preparation; (2) preliminary chromosome fractionation; (3) fractionation of a polystyrene aggregate model which approximates the chromosome shape. We have been successful in isolating metaphase chromosomes from the Chinese hamster. We also received a human chromosome sample from Dr. Carolyn Bell-Prince of Los Alamos National Laboratory. Results are discussed. 2 figs.

  18. A Critical Review of Concepts and Methods Used in Classical Genome Analysis

    DEFF Research Database (Denmark)

    Seberg, Ole; Petersen, Gitte

    1998-01-01

    A short account of the development of classical genome analysis, the analysis of chromosome behaviour in metaphase I of meiosis, primarily in interspecific hybrids, is given. The application of the concept of homology to describe chromosome pairing between the respective chromosomes of a pair...... breeding but it has no place in systematics. With an increased knowledge and understanding of the mechanism behind meiosis, data useful in a systematic context may eventually be produced....

  19. Effects of x-rays on growth of plants and mitotic chromosomal aberrations of Lathyrus sativus Linn

    International Nuclear Information System (INIS)

    Chaudhuri, D.; Das, A.

    1985-01-01

    It has been found that the abnormalities of chromosome at different stages of mitosis show a linear dose relationship. From the detailed study of normal, abnormal phases of prophase, metaphase, anaphase and telophase, it is observed that the abnormality (per cent) in all stages of mitosis has increased with increase in dose. Under different doses, the observed characters of abnormality in chromosomes of Lathyrus sativus may exhibit the occurence of direct hit process. (M.N.)

  20. Mosaic 22q11.2 microdeletion syndrome: diagnosis and clinical manifestations of two cases

    OpenAIRE

    Kabra Madhulika; Jain Manish; Halder Ashutosh; Gupta Neerja

    2008-01-01

    Abstract Chromosome 22q11.2 microdeletion syndrome is due to microdeletion of 22q11.2 region of chromosome 22. It is a common microdeletion syndrome however mosaic cases are very rare and reported only few previous occasions. In this report we describe two unrelated male children with clinical features consistent with 22q11.2 microdeletion syndrome characterized by cardiac defect, facial dysmorphism and developmental deficiency. One of the cases also had trigonocephaly. Interphase & metaphase...

  1. PTP-S2, a nuclear tyrosine phosphatase, is phosphorylated and ...

    Indian Academy of Sciences (India)

    2.1 Cell lines. Rat fibroblast cell line F-111 and D3, a stable clone of HeLa cells expressing rat PTP-S2, were maintained in DMEM containing 10% fetal bovine serum. D3 cell line has been described previously (Radha et al 1997). Cells in exponential cultures were used as interphase cells. Metaphase-arrested cells were ...

  2. Chromosome painting for plant biotechnology.

    Science.gov (United States)

    Kato, Akio; Lamb, Jonathan C; Albert, Patrice S; Danilova, Tatiana; Han, Fangpu; Gao, Zhi; Findley, Seth; Birchler, James A

    2011-01-01

    Fluorescence in situ hybridization (FISH) is an invaluable tool for chromosome analysis and engineering. The ability to visually localize endogenous genes, transposable elements, transgenes, naturally occurring organellar DNA insertions - essentially any unique sequence larger than 2 kb - greatly facilitates progress. This chapter details the labeling procedures and chromosome preparation techniques used to produce high-quality FISH signals on somatic metaphase and meiotic pachytene spreads.

  3. Biological dosimetry by the triage dicentric chromosome assay - Further validation of international networking

    Energy Technology Data Exchange (ETDEWEB)

    Wilkins, Ruth C., E-mail: Ruth.Wilkins@hc-sc.gc.ca [Health Canada, Ottawa, ON K1A 0K9 (Canada); Romm, Horst; Oestreicher, Ursula [Bundesamt fur Strahlenschutz, 38226 Salzgitter (Germany); Marro, Leonora [Health Canada, Ottawa, ON K1A 0K9 (Canada); Yoshida, Mitsuaki A. [Biological Dosimetry Section, Dept. of Dose Assessment, Research Center for Radiation Emergency Medicine, NIRS, 4-9-1 Anagawa, Inage-ku, Chiba 263-8555 (Japan); Department Radiation Biology, Institute of Radiation Emergency Medicine, Hirosaki University Graduate School of Health Sciences, 66-1 Hon-cho, Hirosaki, Aomori 036-8564 (Japan); Suto, Y. [Biological Dosimetry Section, Dept. of Dose Assessment, Research Center for Radiation Emergency Medicine, NIRS, 4-9-1 Anagawa, Inage-ku, Chiba 263-8555 (Japan); Prasanna, Pataje G.S. [National Cancer Institute, Division of Cancer Treatment and Diagnosis, Radiation Research Program, 6130 Executive Blvd., MSC 7440, Bethesda, MD 20892-7440 (United States)

    2011-09-15

    Biological dosimetry is an essential tool for estimating radiation doses received to personnel when physical dosimetry is not available or inadequate. The current preferred biodosimetry method is based on the measurement of radiation-specific dicentric chromosomes in exposed individuals' peripheral blood lymphocytes. However, this method is labor-, time- and expertise-demanding. Consequently, for mass casualty applications, strategies have been developed to increase its throughput. One such strategy is to develop validated cytogenetic biodosimetry laboratory networks, both national and international. In a previous study, the dicentric chromosome assay (DCA) was validated in our cytogenetic biodosimetry network involving five geographically dispersed laboratories. A complementary strategy to further enhance the throughput of the DCA among inter-laboratory networks is to use a triage DCA where dose assessments are made by truncating the labor-demanding and time-consuming metaphase spread analysis to 20 - 50 metaphase spreads instead of routine 500 - 1000 metaphase spread analysis. Our laboratory network also validated this triage DCA, however, these dose estimates were made using calibration curves generated in each laboratory from the blood samples irradiated in a single laboratory. In an emergency situation, dose estimates made using pre-existing calibration curves which may vary according to radiation type and dose rate and therefore influence the assessed dose. Here, we analyze the effect of using a pre-existing calibration curve on assessed dose among our network laboratories. The dose estimates were made by analyzing 1000 metaphase spreads as well as triage quality scoring and compared to actual physical doses applied to the samples for validation. The dose estimates in the laboratory partners were in good agreement with the applied physical doses and determined to be adequate for guidance in the treatment of acute radiation syndrome.

  4. Impact of various parameters in detecting chromosomal aberrations by FISH to describe radiosensitivity

    International Nuclear Information System (INIS)

    Keller, U.; Mueller, E.; Grabenbauer, G.; Sauer, R.; Distel, L.; Kuechler, A.; Liehr, T.

    2004-01-01

    Background and purpose: analysis of radiation-induced chromosomal aberrations is regarded as the ''gold standard'' for classifying individual radiosensitivity. A variety of different parameters can be used. The crucial question, however, is to explore which parameter is suited best to describe the differences between patients with increased radiosensitivity and healthy individuals. Patients and methods: in this study, five patients with severe radiation-induced late effects of at least grade 3, classified according to the Radiation Therapy Oncology Group (RTOG), and eleven healthy individuals were examined retrospectively. Peripheral blood lymphocytes were irradiated in vitro with 0.7 Gy and 2.0 Gy prior to cultivation and stained by means of three-color fluorescence in situ hybridization (FISH). The detailed analysis was focused on the number of breaks per metaphase, on breaks from complex chromosomal rearrangements per metaphase, as well as on the percentage of translocations, dicentric chromosomes, breaks, and excess acentric fragments - each in comparison with the total number of mitoses analyzed. Results: using the number of breaks from complex chromosomal rearrangements after 2.0 Gy, radiosensitive patients as endpoint were clearly to be distinguished (p = 0.001) from healthy individuals. Translocations (p = 0.001) as well as breaks per metaphase (p = 0.002) were also suitable indicators for detecting differences between patients and healthy individuals. The parameters ''percentage of dicentric chromosomes'', ''breaks'', and ''excess acentric fragments'' in comparison to the total number of mitoses analyzed could neither serve as meaningful nor as significant criteria, since they showed a strong interindividual variability. Conclusion: to detect a difference in chromosomal aberrations between healthy and radiosensitive individuals, the parameters ''frequency of breaks per metaphase'', ''complex chromosomal rearrangements'', and ''translocations'' are most

  5. Epigenetic inactivation of CHFR in human tumors

    OpenAIRE

    Toyota, Minoru; Sasaki, Yasushi; Satoh, Ayumi; Ogi, Kazuhiro; Kikuchi, Takefumi; Suzuki, Hiromu; Mita, Hiroaki; Tanaka, Nobuyuki; Itoh, Fumio; Issa, Jean-Pierre J.; Jair, Kam-Wing; Schuebel, Kornel E.; Imai, Kohzoh; Tokino, Takashi

    2003-01-01

    Cell-cycle checkpoints controlling the orderly progression through mitosis are frequently disrupted in human cancers. One such checkpoint, entry into metaphase, is regulated by the CHFR gene encoding a protein possessing forkhead-associated and RING finger domains as well as ubiquitin–ligase activity. Although defects in this checkpoint have been described, the molecular basis and prevalence of CHFR inactivation in human tumors are still not fully understood. To address this question, w...

  6. Detection of illegitimate rearrangement within the immunoglobulin locus on 14q32.3 in B-cell malignancies using end-sequenced probes

    DEFF Research Database (Denmark)

    Silahtaroglu, Asli; Poulsen, Tim; Giselø, Claus

    2001-01-01

    in the literature or at http://www.biologia.uniba.it/rmc/. Validation, orientation, and overlap of these probes were confirmed using interphase-, metaphase-, and fiber-FISH. We have identified seven BAC end-sequenced probes (3087C18, 47P23, 76N15, 12F16, 101G24, 112H5, and 151B17) covering 612 kb of the distal IGHV...

  7. In situ hybridization at the electron microscope level: hybrid detection by autoradiography and colloidal gold.

    Science.gov (United States)

    Hutchison, N J; Langer-Safer, P R; Ward, D C; Hamkalo, B A

    1982-11-01

    In situ hybridization has become a standard method for localizing DNA or RNA sequences in cytological preparations. We developed two methods to extend this technique to the transmission electron microscope level using mouse satellite DNA hybridization to whole mount metaphase chromosomes as the test system. The first method devised is a direct extension of standard light microscope level using mouse satellite DNA hybridization to whole mount metaphase chromosomes as the test system. The first method devised is a direct extension of standard light microscope in situ hybridization. Radioactively labeled complementary RNA (cRNA) is hybridized to metaphase chromosomes deposited on electron microscope grids and fixed in 70 percent ethanol vapor; hybridixation site are detected by autoradiography. Specific and intense labeling of chromosomal centromeric regions is observed even after relatively short exposure times. Inerphase nuclei present in some of the metaphase chromosome preparations also show defined paatterms of satellite DNA labeling which suggests that satellite-containing regions are associate with each other during interphase. The sensitivity of this method is estimated to at least as good as that at the light microscope level while the resolution is improved at least threefold. The second method, which circumvents the use of autoradiogrphic detection, uses biotin-labeled polynucleotide probes. After hybridization of these probes, either DNA or RNA, to fixed chromosomes on grids, hybrids are detected via reaction is improved at least threefold. The second method, which circumvents the use of autoradiographic detection, uses biotin-labeled polynucleotide probes. After hybridization of these probes, either DNA or RNA, to fixed chromosomes on grids, hybrids are detected via reaction with an antibody against biotin and secondary antibody adsorbed to the surface of over centromeric heterochromatin and along the associated peripheral fibers. Labeling is on average

  8. Przebieg mitozy w korzeniach siewek pszenicy pod wpływem rubidu [Course of mitosis in root seedlings of Triticum aestivum L. under the influence of rubidium

    Directory of Open Access Journals (Sweden)

    Kazimierz Olech

    2015-06-01

    Full Text Available The depresion of mitosis as a result of rubidium action illustrated by the lower frequency of mitosis was found in an investigation made on roots of wheat seedlings grown on a nutrient medium which the total amount of potassium or a half of it was replaced by rubidium. The lower mitotic activity was caused by limited number of nuclei beginning prophase and by a prolonged duration of metaphase.

  9. Role of the pseudoautosomal region in sex-chromosome pairing during male meiosis: Meiotic studies in a man with a deletion of distal Xp

    Energy Technology Data Exchange (ETDEWEB)

    Mohandas, T.K.; Passage, M.B.; Yen, P.H.; Speed, R.M.; Chandley, A.C.; Shapiro, L.J. (Univ. of California, San Francisco, CA (United States))

    1992-09-01

    Meiotic studies were undertaken in a 24-year-old male patient with short stature, chondrodysplasia punctata, ichthyosis, steroid sulfatase deficiency, and mild mental retardation with an inherited cytologically visible deletion of distal Xp. Molecular investigations showed that the pseudoautosomal region as well as the steroid sulfatase gene were deleted, but telomeric sequences were present at the pter on the deleted X chromosome. A complete failure of sex-chromosome pairing was observed in the primary spermatocytes of the patient. Telomeric approaches between the sex chromosomes were made at zygotene in some cells, but XY synaptonemal complex was formed. The sex chromosomes were present as univalents at metaphase I, and germ-cell development was arrested between metaphase I and metaphase II in the vast majority of cells, consistent with the azoospermia observed in the patient. The failure of XY pairing in this individual indicates that the pseudoautosomal sequences play an important role in initiating XY pairing and formation of synaptonemal complex at meiosis. 36 refs., 6 figs.

  10. Premature centromere division and other centromeric misbehavior

    Energy Technology Data Exchange (ETDEWEB)

    Fitzgerald, P.H. [Christchurch School of Medicine (New Zealand)

    1993-12-31

    Premature centromere division was initially described for the X chromosome. In an otherwise typical metaphase cell, one chromosome showed no primary constriction and appeared to have no centromere. G-banding analysis indicated that this apparent acentric fragment was an entire X chromosome. Because its centromere was divided when the centromeres of all other chromosomes of the metaphase cell were entire, the condition was described as premature centromere division (PCD). The importance of PCD lies in its being a mechanism on non-disjunction, as was indicated by the strong association of X chromosome aneuploidy with PCD,X. We can infer that the affected chromosome failed to take part in the normal distribution of chromosomes at mitoses. The centromere, it its widest sense, is generally believed to have a role in the correct orientation of chromosomes at the metaphase plate and the distribution of chromatids to the spindle poles. The failure of these functions implies a major centromeric dysfunction. What do we know of this complex region of the chromosome that might help us understand its dysfunction?

  11. Genotoxicity and cytotoxicity assessment of new ethyl-carbamates with ixodicidal activity.

    Science.gov (United States)

    Prado-Ochoa, María Guadalupe; Muñoz-Guzmán, Marco Antonio; Vázquez-Valadez, Víctor Hugo; Velázquez-Sánchez, Ana María; Salazar, Ana María; Ramírez-Noguera, Patricia; Angeles, Enrique; Alba-Hurtado, Fernando

    2016-09-01

    The mammalian erythrocyte micronucleus test was used on the peripheral blood of Wistar rats exposed to two new ethyl-carbamates: ethyl-4-bromophenyl-carbamate (LQM 919) and ethyl-4-chlorophenyl-carbamate (LQM 996) to analyze their genotoxic potential. The mitotic index and cell proliferation kinetics in human lymphocyte cultures in the presence of these ethyl-carbamates were used to evaluate cytotoxicity and cytostaticity respectively. Exposure to greater acute doses (300mg/kg) and to all of the subchronic doses (12.5, 25 and 50mg/kg daily for 90 days) of these ethyl-carbamates induced an increased frequency (pcarbamates. Increases in MN-PCE was higher in males than in females exposed to LQM 996 50mg/Kg (pcarbamate exposure. The highest concentration (0.3mM) of both ethyl-carbamates in lymphocyte cultures increased the percentage of cells in first division metaphase and decreased the percentage of cells in third division metaphase, indicating an increase in cell cycle length or a possible cell cycle arrest in metaphase (cytostatic effect). The results of this study show that the evaluated ethyl-carbamates may induce genotoxic damage in rats and alterations in the human lymphocyte cell cycle. Copyright © 2016 Elsevier B.V. All rights reserved.

  12. Quantitative analysis of comparative genomic hybridization

    Energy Technology Data Exchange (ETDEWEB)

    Manoir, S. du; Bentz, M.; Joos, S. [Abteilung Organisation komplexer Genome, Heidelberg (Germany)]|[Institut fuer Humangenetik, Heidelberg (Germany)] [and others

    1995-01-01

    Comparative genomic hybridization (CGH) is a new molecular cytogenetic method for the detection of chromosomal imbalances. Following cohybridization of DNA prepared from a sample to be studied and control DNA to normal metaphase spreads, probes are detected via different fluorochromes. The ratio of the test and control fluorescence intensities along a chromosome reflects the relative copy number of segments of a chromosome in the test genome. Quantitative evaluation of CGH experiments is required for the determination of low copy changes, e.g., monosomy or trisomy, and for the definition of the breakpoints involved in unbalanced rearrangements. In this study, a program for quantitation of CGH preparations is presented. This program is based on the extraction of the fluorescence ratio profile along each chromosome, followed by averaging of individual profiles from several metaphase spreads. Objective parameters critical for quantitative evaluations were tested, and the criteria for selection of suitable CGH preparations are described. The granularity of the chromosome painting and the regional inhomogeneity of fluorescence intensities in metaphase spreads proved to be crucial parameters. The coefficient of variation of the ratio value for chromosomes in balanced state (CVBS) provides a general quality criterion for CGH experiments. Different cutoff levels (thresholds) of average fluorescence ratio values were compared for their specificity and sensitivity with regard to the detection of chromosomal imbalances. 27 refs., 15 figs., 1 tab.

  13. Prophase I Mouse Oocytes Are Deficient in the Ability to Respond to Fertilization by Decreasing Membrane Receptivity to Sperm and Establishing a Membrane Block to Polyspermy1

    Science.gov (United States)

    Kryzak, Cassie A.; Moraine, Maia M.; Kyle, Diane D.; Lee, Hyo J.; Cubeñas-Potts, Caelin; Robinson, Douglas N.; Evans, Janice P.

    2013-01-01

    ABSTRACT Changes occurring as the prophase I oocyte matures to metaphase II are critical for the acquisition of competence for normal egg activation and early embryogenesis. A prophase I oocyte cannot respond to a fertilizing sperm as a metaphase II egg does, including the ability to prevent polyspermic fertilization. Studies here demonstrate that the competence for the membrane block to polyspermy is deficient in prophase I mouse oocytes. In vitro fertilization experiments using identical insemination conditions result in monospermy in 87% of zona pellucida (ZP)-free metaphase II eggs, while 92% of ZP-free prophase I oocytes have four or more fused sperm. The membrane block is associated with a postfertilization reduction in the capacity to support sperm binding, but this reduction in sperm-binding capacity is both less robust and slower to develop in fertilized prophase I oocytes. Fertilization of oocytes is dependent on the tetraspanin CD9, but little to no release of CD9 from the oocyte membrane is detected, suggesting that release of CD9-containing vesicles is not essential for fertilization. The deficiency in membrane block establishment in prophase I oocytes correlates with abnormalities in two postfertilization cytoskeletal changes: sperm-induced cortical remodeling that results in fertilization cone formation and a postfertilization increase in effective cortical tension. These data indicate that cortical maturation is a component of cytoplasmic maturation during the oocyte-to-egg transition and that the egg cortex has to be appropriately primed and tuned to be responsive to a fertilizing sperm. PMID:23863404

  14. Hamster oocyte membrane potential and ion permeability vary with preantral cumulus cell attachment and developmental stage

    Directory of Open Access Journals (Sweden)

    Miller Raymond L

    2001-10-01

    Full Text Available Abstract Background In vitro maturation of mammalian oocytes is an area of great interest due to its potential application in the treatment of infertility. The morphological and physiological changes that occur during oocyte development are poorly understood, and further studies are needed investigating the physiological changes associated with oocyte maturation. In this study we evaluated the membrane potential and the sodium/potassium permeability ratio of oocytes acutely isolated, and cumulus-oocyte complexes in metaphase II and preantral follicle stages. Results Intracellular electrical recordings revealed that cumulus-enclosed oocytes have a membrane potential significantly more negative at the preantral follicle stage than at metaphase II stage (-38.4 versus -19.7 mV, p Conclusions These data show a change in the membrane potential and Na+/K+ permeability ratio during ooycte development from the preantral stage oocyte to the metaphase II stage. We have also demonstrated a change in the preantral oocyte membrane potential when surrounding cumulus cells are removed; either due to membrane changes or loss of cumulus cells.

  15. Ric-8A and Gi alpha recruit LGN, NuMA, and dynein to the cell cortex to help orient the mitotic spindle.

    Science.gov (United States)

    Woodard, Geoffrey E; Huang, Ning-Na; Cho, Hyeseon; Miki, Toru; Tall, Gregory G; Kehrl, John H

    2010-07-01

    In model organisms, resistance to inhibitors of cholinesterase 8 (Ric-8), a G protein alpha (G alpha) subunit guanine nucleotide exchange factor (GEF), functions to orient mitotic spindles during asymmetric cell divisions; however, whether Ric-8A has any role in mammalian cell division is unknown. We show here that Ric-8A and G alpha(i) function to orient the metaphase mitotic spindle of mammalian adherent cells. During mitosis, Ric-8A localized at the cell cortex, spindle poles, centromeres, central spindle, and midbody. Pertussis toxin proved to be a useful tool in these studies since it blocked the binding of Ric-8A to G alpha(i), thus preventing its GEF activity for G alpha(i). Linking Ric-8A signaling to mammalian cell division, treatment of cells with pertussis toxin, reduction of Ric-8A expression, or decreased G alpha(i) expression similarly affected metaphase cells. Each treatment impaired the localization of LGN (GSPM2), NuMA (microtubule binding nuclear mitotic apparatus protein), and dynein at the metaphase cell cortex and disturbed integrin-dependent mitotic spindle orientation. Live cell imaging of HeLa cells expressing green fluorescent protein-tubulin also revealed that reduced Ric-8A expression prolonged mitosis, caused occasional mitotic arrest, and decreased mitotic spindle movements. These data indicate that Ric-8A signaling leads to assembly of a cortical signaling complex that functions to orient the mitotic spindle.

  16. NuMA phosphorylation by CDK1 couples mitotic progression with cortical dynein function.

    Science.gov (United States)

    Kotak, Sachin; Busso, Coralie; Gönczy, Pierre

    2013-09-11

    Spindle positioning and spindle elongation are critical for proper cell division. In human cells, an evolutionary conserved ternary complex (NuMA/LGN/Gαi) anchors dynein at the cortex during metaphase, thus ensuring correct spindle positioning. Whether this complex contributes to anaphase spindle elongation is not known. More generally, the mechanisms coupling mitotic progression with spindle behaviour remain elusive. Here, we uncover that levels of cortical dynein markedly increase during anaphase in a NuMA-dependent manner. We demonstrate that during metaphase, CDK1-mediated phosphorylation at T2055 negatively regulates NuMA cortical localization and that this phosphorylation is counteracted by PPP2CA phosphatase activity. We establish that this tug of war is essential for proper levels of cortical dynein and thus spindle positioning during metaphase. Moreover, we find that upon CDK1 inactivation in anaphase, the rise in dephosphorylated NuMA at the cell cortex leads to cortical dynein enrichment, and thus to robust spindle elongation. Our findings uncover a mechanism whereby the status of NuMA phosphorylation coordinates mitotic progression with proper spindle function.

  17. Spindle pole cohesion requires glycosylation-mediated localization of NuMA.

    Science.gov (United States)

    Magescas, Jérémy; Sengmanivong, Lucie; Viau, Amandine; Mayeux, Adeline; Dang, Tien; Burtin, Martine; Nilsson, Ulf J; Leffler, Hakon; Poirier, Françoise; Terzi, Fabiola; Delacour, Delphine

    2017-05-03

    Glycosylation is critical for the regulation of several cellular processes. One glycosylation pathway, the unusual O-linked β-N-acetylglucosamine glycosylation (O-GlcNAcylation) has been shown to be required for proper mitosis, likely through a subset of proteins that are O-GlcNAcylated during metaphase. As lectins bind glycosylated proteins, we asked if specific lectins interact with mitotic O-GlcNAcylated proteins during metaphase to ensure correct cell division. Galectin-3, a small soluble lectin of the Galectin family, is an excellent candidate, as it has been previously described as a transient centrosomal component in interphase and mitotic epithelial cells. In addition, it has recently been shown to associate with basal bodies in motile cilia, where it stabilizes the microtubule-organizing center (MTOC). Using an experimental mouse model of chronic kidney disease and human epithelial cell lines, we investigate the role of Galectin-3 in dividing epithelial cells. Here we find that Galectin-3 is essential for metaphase where it associates with NuMA in an O-GlcNAcylation-dependent manner. We provide evidence that the NuMA-Galectin-3 interaction is important for mitotic spindle cohesion and for stable NuMA localization to the spindle pole, thus revealing that Galectin-3 is a novel contributor to epithelial mitotic progress.

  18. Genotoxic effects of borax on cultured lymphocytes.

    Science.gov (United States)

    Pongsavee, Malinee

    2009-03-01

    The effect of borax on human chromosomes was analyzed in this study. Venous blood from 30 male students at Thammasat University, Thailand (age 18-25 years) was collected for lymphocyte cell cultures. This experiment was divided into two groups: the first group was the control group and the second group was the experimental group. The lymphocyte cells in the control group were cultured without borax. The experimental group was divided into four subgroups. The lymphocyte cells in each experimental subgroup were cultured with different concentrations of borax (0.1 mg/ml, 0.15 mg/ml, 0.2 mg/ml and 0.3 mg/ml). Human chromosomes were studied for abnormalities through Giemsa-staining and G-banding. The results show that the numbers of metaphase plates (the metaphase plate which contained 46 chromosomes; 46, XY) and metaphase chromosomes were reduced when lymphocyte cells were cultured with 0.15 mg/ml (57.2%), 0.2 mg/ml (50.8%) and 0.3 mg/ml (42.3%) concentrations of borax. There was a statistically significant difference between the control and experimental subgroups (p borax concentration experimental subgroup. This shows that borax (at 0.15, 0.2 and 0.3 mg/ml concentrations) affects the cell and human chromosomes (both numerical and structural abnormalities). Borax may cause human chromosome abnormalities and lead to genetic defects.

  19. Carbamazepine induces mitotic arrest in mammalian Vero cells

    International Nuclear Information System (INIS)

    Perez Martin, J.M.; Fernandez Freire, P.; Labrador, V.; Hazen, M.J.

    2008-01-01

    We reported recently that the anticonvulsant drug carbamazepine, at supratherapeutic concentrations, exerts antiproliferative effects in mammalian Vero cells, but the underlying mechanism has not been elucidated. This motivates us to examine rigorously whether growth arrest was associated with structural changes in cellular organization during mitosis. In the present work, we found that exposure of the cells to carbamazepine led to an increase in mitotic index, mainly due to the sustained block at the metaphase/anaphase boundary, with the consequent inhibition of cell proliferation. Indirect immunofluorescence, using antibodies directed against spindle apparatus proteins, revealed that mitotic arrest was associated with formation of monopolar spindles, caused by impairment of centrosome separation. The final consequence of the spindle defects induced by carbamazepine, depended on the duration of cell cycle arrest. Following the time course of accumulation of metaphase and apoptotic cells during carbamazepine treatments, we observed a causative relationship between mitotic arrest and induction of cell death. Conversely, cells released from the block of metaphase by removal of the drug, continued to progress through mitosis and resume normal proliferation. Our results show that carbamazepine shares a common antiproliferative mechanism with spindle-targeted drugs and contribute to a better understanding of the cytostatic activity previously described in Vero cells. Additional studies are in progress to extend these initial findings that define a novel mode of action of carbamazepine in cultured mammalian cells

  20. Cytosolic Cl- Affects the Anticancer Activity of Paclitaxel in the Gastric Cancer Cell Line, MKN28 Cell

    Directory of Open Access Journals (Sweden)

    Sachie Tanaka

    2017-05-01

    Full Text Available Background/Aims: Our previous study revealed that cytosolic Cl- affected neurite elongation promoted via assembly of microtubule in rat pheochromocytoma PC12D cells and Cl-–induced blockade of intrinsic GTPase enhanced tubulin polymerization in vitro. Paclitaxel (PTX is a microtubule-targeted chemotherapeutic drug and stabilizes microtubules resulting in mainly blockade of mitosis at the metaphase-anaphase transition and induction of apoptosis. In the present study, we tried to clarify whether the cytosolic Cl- affected PTX ability to inhibit cell growth in the gastric cancer cell line, MKN28. Methods: To clarify the cytosolic Cl- action on PTX-induced cell death and metaphase-anaphase transition in the gastric cancer cell line, MKN28 cell, and PTX-induced tubulin polymerization, we performed cell proliferation assay, cytosolic Cl- concentration measurement, immunofluorescence microscopy, and in vitro tubulin polymerization assay. Results: The decline of cytosolic Cl- weakened the cytotoxic effect of PTX on cell proliferation of MKN28 cells, which could pass through the metaphase-anaphase transition. Moreover, in vitro PTX-induced tubulin polymerization was diminished under the low Cl- condition. Conclusions: Our results strongly suggest that the upregulation of cytosolic Cl- concentration would enhance the antitumor effect of PTX, and that the cytosolic Cl- would be one of the key targets for anti-cancer therapy.

  1. The first cytogenetic characterization of the poisonous black widow spider Latrodectus gr. curacaviensis from Brazil, with chromosomal review of the family Theridiidae (Arachnida, Araneae).

    Science.gov (United States)

    Araujo, Douglas; Maia, Ulysses Madureira; Brescovit, Antonio Domingos

    2010-02-01

    In this paper we present, for the first time, cytogenetical data on Latrodectus gr. curacaviensis (Theridiidae) from Brazil, as well as the first data on meiosis and sex chromosome system of this genus. Testes were submitted to colchicine, hypotonic, and fixation treatment, and chromosomal preparations were stained with Giemsa solution. The analysis showed 2n=26 telo/acrocentric chromosomes in spermatogonial metaphases. Metaphase I exhibited 12 autosomal bivalents and two sex chromosome univalents (12II + X(1)X(2)). All bivalents revealed one terminal chiasma. Metaphases II confirmed the sex chromosome system, showing 12 autosomes or 12 autosomes plus two X chromosomes, respectively. Male karyotype prevailing in theridiids is formed by 2n=22 chromosomes, including sex chromosome system X(1)X(2) in all species. The Latrodectus species of the geometricus clade analyzed until now showed smaller diploid number (2nfemale symbol=16 and 2nfemale symbol=18) than the species of the mactans clade (2nfemale symbol=24 and 2nfemale symbol=26). Thus, according to the chromosome number, the examined Latrodectus species seems to be related to the mactans clade.

  2. Measurement of background translocation frequencies in individuals with clones

    Energy Technology Data Exchange (ETDEWEB)

    Wade, Marcelle J. [California State Univ. (CalState), Hayward, CA (United States)

    1996-08-01

    In the leukemia case the unseparated B and T lymphocytes had a high translocation frequency even after 0.0014, respectively. After purging all clones from the data, the translocation frequencies for Bio 8 and Bio 23 were 0.00750.0014 and 0.0073 metaphases were scored for chromosomal aberrations,, specifically reciprocal translocations, using fluorescence in situ hybridization (FISH). Metaphase spreads were used from two healthy, unexposed individuals (not exposed to radiation, chemotherapy or radiotherapy) and one early B- precursor acute lymphocytic leukemia (ALL) patient (metaphase spreads from both separated T lymphocytes and unseparated B and T lymphocytes were scored). All three individuals had an abnormally high translocation frequency. The high translocation frequencies resulted from clonal expansion of specific translocated chromosomes. I show in this thesis that by purging (discounting or removing) clones from the data of unexposed individuals, one can obtain true background translocation frequencies. In two cases, Bio 8 and Bio 23, the measured translocation frequency for chromosomes 1, 2 and 4 was 0.0124 purging all of the clones from the data. This high translocation frequency may be due to a low frequency of some clones and may not be recognized. The separated T lymphocytes had a higher translocation frequency than expected.

  3. APC-targeted RAA1 degradation mediates the cell cycle and root development in plants.

    Science.gov (United States)

    Xu, Yunyuan; Cao, Hong; Chong, Kang

    2010-03-01

    Protein degradation by the ubiquitin-proteasome system is necessary for a normal cell cycle. As compared with knowledge of the mechanism in animals and yeast, that in plants is less known. Here we summarize research into the regulatory mechanism of protein degradation in the cell cycle in plants. Anaphase-promoting complex/cyclosome (APC), in the E3 family of enzymes, plays an important role in maintaining normal mitosis. APC activation and substrate specificity is determined by its activators, which can recognize the destruction box (D-box) in APC target proteins. Oryza sativa root architecture-associated 1 (OsRAA1) with GTP-binding activity was originally cloned from rice. Overexpression of of OsRAA1 inhibits the growth of primary roots in rice. Knockdown lines showed reduced height of seedlings because of abnormal cell division. OsRAA1 transgenic rice and fission yeast show a higher proportion of metaphase cells than that of controls, which suggests a blocked transition from metaphase to anaphase during mitosis. OsRAA1 co-localizes with spindle tubulin. It contains the D-box motif and interacts with OsRPT4 of the regulatory particle of 26S proteasome. OsRAA1 may be a cell cycle inhibitor that can be degraded by the ubiquitin-proteasome system, and its disruption is necessary for the transition from metaphase to anaphase during root growth in rice.

  4. APC-targeted RAAI degradation mediates the cell cycle and root development in plants

    Science.gov (United States)

    Cao, Hong; Chong, Kang

    2010-01-01

    Protein degradation by the ubiquitin-proteasome system is necessary for a normal cell cycle. As compared with knowledge of the mechanism in animals and yeast, that in plants is less known. Here we summarize research into the regulatory mechanism of protein degradation in the cell cycle in plants. Anaphase-promoting complex/cyclosome (APC), in the E3 family of enzymes, plays an important role in maintaining normal mitosis. APC activation and substrate specificity is determined by its activators, which can recognize the destruction box (D-box) in APC target proteins. Oryza sativa root architecture-associated I (OsRAA1) with GTP-binding activity was originally cloned from rice. Overexpression of of OsRAA1 inhibits the growth of primary roots in rice. Knockdown lines showed reduced height of seedlings because of abnormal cell division. OsRAA1 transgenic rice and fission yeast show a higher proportion of metaphase cells than that of controls, which suggests a blocked transition from metaphase to anaphase during mitosis. OsRAA1 co-localizes with spindle tubulin. It contains the D-box motif and interacts with OsRPT4 of the regulatory particle of 26S proteasome. OsRAA1 may be a cell cycle inhibitor that can be degraded by the ubiquitin-proteasome system, and its disruption is necessary for the transition from metaphase to anaphase during root growth in rice. PMID:20037474

  5. Automatic aberration scoring using whole chromosome F.I.S.H

    International Nuclear Information System (INIS)

    Piper, J.; Bayley, R.; Boyle, S.; Fantes, J.A.; Green, D.K.; Gordon, J.; Hill, W.; Ji, L.; Malloy, P.; Perry, P.; Rutovitz, D.; Stark, M.; Whale, D.

    1993-01-01

    A radiation-induced rearrangement involving a painted and a non-painted chromosome will usually result in two partly-painted chromosomes, typically either a dicentric chromosome and associated fragment, or a reciprocal translocation pair. A consequence of such a rearrangement is that the number of painted image regions in the metaphase is increased by one, and their size distribution is altered. More complex rearrangements are uncommon, particularly at low doses. A high proportion of damaged cells can therefore be registered simply by detecting when the distribution of painted components differs from the expected number and size. A system has been constructed to pre-screen for damaged cells. It comprises automatic fluorescence metaphase finding followed by relocation and digitization of probe and counterstain channels at high resolution. Fully automatic segmentation in counterstain discriminates chromosomes from interphase nuclei and determines whether a metaphase is approximately diploid. The painted regions are segmented and their relative sizes estimated. Rules are applied which reduce the false positives due to artifacts such as overlapped painted chromosomes. More than 70% of cells with radiation damage involving painted and unpainted chromosomes were detected in a preliminary experiment using a small data set, with a low false positive rate. Results from a larger experiment in progress are presented

  6. C-banding and fluorescent in situ hybridization with rDNA sequences in chromosomes of Cycloneda sanguinea Linnaeus (Coleoptera, Coccinellidae

    Directory of Open Access Journals (Sweden)

    Eliane Mariza Dortas Maffei

    2004-01-01

    Full Text Available The aim of this study was to describe mitotic and meiotic chromosomes of Cycloneda sanguinea using C-banding, fluorescent in situ hybridization (FISH rDNA probes, and sequential FISH/Ag-NOR staining. The chromosome number was 2n = 18 + XX for females and 2n = 18 + Xy for males. The X chromosome was metacentric and the Y chromosome was very small. During meiosis, the karyotypic meioformula was n = 9 + Xy p, and sex chromosomes configured a parachute at metaphase I. At the beginning of pachytene, bivalents were still individualized, and sex chromosomes were associated end-to-end through the heteropycnotic region of the X chromosome. Later in pachytene, further condensation led to the formation of a pseudo-ring by the sex bivalent. All chromosomes showed pericentromeric heterochromatin. FISH and sequential FISH/Ag-NOR staining evidenced the location of the nucleolar organizer region in one pair of autosomes (at spermatogonial metaphase. During meiosis, these genes were mapped to a region outside the sex vesicle by FISH, although Xy p was deeply stained with silver at metaphase I. These results suggest that these argyrophilic substances are of a nucleolar protein nature, and seem to be synthesized by a pair of autosomes and imported during meiosis (prophase I to the sex pair, during the association of the sex chromosomes.

  7. Cytogenetic studies with laser or X-ray exposures

    International Nuclear Information System (INIS)

    Bozduganov, A.; Genkov, P.

    1975-01-01

    Account is given of studies involving a total of 21 peripheral blood cultures given the following treatments: exposure to 20 0.13-joule pulses from an optic quantum generator (ruby), 9; exposure to 1000 R X-rays, 9; and unexposed controls, 3. Exposures were carried out on three cultures from each experimental series at each of three time intervals, namely 24 h, 48 h, and 72 h after initiation of cultures. On any day, 40 well-spread metaphases were examined. High quality metaphases were photographed and karyotyped in conformity to the Chicago Conference criteria. In the laser experiment, chromosome aberrations were observed in 65% of methaphases analyzed vs. 5.3% in controls. Anomalies encountered included aneuploidy, with hypoploid metaphases predominating, polyploidy (triploidy, tetraploidy, and partial endoreduplication), and structural alterations. The following structural chromosome rearrangements are found: acentric fragments, mostly pairs, occasionally single, including minute chromosomes; dicentric and tricentric chromosomes; interstitial deletions; and chromosome translocations. Most varied and abundant chromosome aberrations were seen in 72-h irradiated cultures. The data presented offer a new opportunity for assessing genetic lesions after laser exposure and may help to determine threshold doses. (author)

  8. Histone-DNA contacts in structure/function relationships of nucleosomes as revealed by crosslinking

    Energy Technology Data Exchange (ETDEWEB)

    Usachenko, S.I. [Univ. of California, Davis, CA (United States); Bradbury, E.M. [Los Alamos National Lab., NM (United States). Life Science Div.]|[Univ. of California, Davis, CA (United States)

    1998-12-31

    The magnitude of the problem of understanding the structure/function relationships of eukaryotic chromosomes can be appreciated from the fact that the human diploid genome contains more than 2 meters of DNA packaged into 46 chromosomes, each at metaphase being several microns in length. Each chromatid of a chromosome contains a single DNA molecule several centimeters in length. In addition to the DNA, chromosomes contain an equal weight of histones and an equal weight of non-histone chromosomal proteins. These histones are the major chromosomal structural proteins. The non-histone chromosomal proteins are involved in the DNA processes of transcription and replication, in chromosome organization and in nuclear architecture. Polytene chromosomes with their bands and interbands and puffs of active genetic loci provide visual evidence for long range order as do the bands and interbands of mammalian metaphase chromosomes. The gentle removal of histones and all but the most tightly bound 2--3% of non-histone proteins from metaphase chromosomes revealed by electron microscopy a residual protein scaffold constraining a halo of DNA loops extending out from the scaffold.

  9. Trans-generational radiation-induced chromosomal instability in the female enhances the action of chemical mutagens

    International Nuclear Information System (INIS)

    Camats, Nuria; Garcia, Francisca; Parrilla, Juan Jose; Calaf, Joaquim; Martin, Miguel; Caldes, Montserrat Garcia

    2008-01-01

    Genomic instability can be produced by ionising radiation, so-called radiation-induced genomic instability, and chemical mutagens. Radiation-induced genomic instability occurs in both germinal and somatic cells and also in the offspring of irradiated individuals, and it is characterised by genetic changes including chromosomal rearrangements. The majority of studies of trans-generational, radiation-induced genomic instability have been described in the male germ line, whereas the authors who have chosen the female as a model are scarce. The aim of this work is to find out the radiation-induced effects in the foetal offspring of X-ray-treated female rats and, at the same time, the possible impact of this radiation-induced genomic instability on the action of a chemical mutagen. In order to achieve both goals, the quantity and quality of chromosomal damage were analysed. In order to detect trans-generational genomic instability, a total of 4806 metaphases from foetal tissues from the foetal offspring of X-irradiated female rats (5 Gy, acute dose) were analysed. The study's results showed that there is radiation-induced genomic instability: the number of aberrant metaphases and the breaks per total metaphases studied increased and were found to be statistically significant (p ≤ 0.05), with regard to the control group. In order to identify how this trans-generational, radiation-induced chromosomal instability could influence the chromosomal behaviour of the offspring of irradiated rat females in front of a chemical agent (aphidicolin), a total of 2481 metaphases were studied. The observed results showed that there is an enhancement of the action of the chemical agent: chromosomal breaks per aberrant metaphases show significant differences (p ≤ 0.05) in the X-ray- and aphidicolin-treated group as regards the aphidicolin-treated group. In conclusion, our findings indicate that there is trans-generational, radiation-induced chromosomal instability in the foetal cells

  10. Trans-generational radiation-induced chromosomal instability in the female enhances the action of chemical mutagens

    Energy Technology Data Exchange (ETDEWEB)

    Camats, Nuria [Institut de Biotecnologia i Biomedicina (IBB), Universitat Autonoma de Barcelona, 08193 Barcelona (Spain); Departament de Biologia Cel.lular, Fisiologia i Immunologia, Universitat Autonoma de Barcelona, 08193 Barcelona (Spain); Garcia, Francisca [Institut de Biotecnologia i Biomedicina (IBB), Universitat Autonoma de Barcelona, 08193 Barcelona (Spain); Parrilla, Juan Jose [Servicio de Ginecologia y Obstetricia, Hospital Universitario Virgen de la Arrixaca, 30120 El Palmar, Murcia (Spain); Calaf, Joaquim [Servei de Ginecologia i Obstetricia, Hospital Universitari de la Santa Creu i Sant Pau, 08025 Barcelona (Spain); Martin, Miguel [Departament de Pediatria, d' Obstetricia i Ginecologia i de Medicina Preventiva, Universitat Autonoma de Barcelona, 08193 Barcelona (Spain); Caldes, Montserrat Garcia [Institut de Biotecnologia i Biomedicina (IBB), Universitat Autonoma de Barcelona, 08193 Barcelona (Spain); Departament de Biologia Cel.lular, Fisiologia i Immunologia, Universitat Autonoma de Barcelona, 08193 Barcelona (Spain)], E-mail: Montserrat.Garcia.Caldes@uab.es

    2008-04-02

    Genomic instability can be produced by ionising radiation, so-called radiation-induced genomic instability, and chemical mutagens. Radiation-induced genomic instability occurs in both germinal and somatic cells and also in the offspring of irradiated individuals, and it is characterised by genetic changes including chromosomal rearrangements. The majority of studies of trans-generational, radiation-induced genomic instability have been described in the male germ line, whereas the authors who have chosen the female as a model are scarce. The aim of this work is to find out the radiation-induced effects in the foetal offspring of X-ray-treated female rats and, at the same time, the possible impact of this radiation-induced genomic instability on the action of a chemical mutagen. In order to achieve both goals, the quantity and quality of chromosomal damage were analysed. In order to detect trans-generational genomic instability, a total of 4806 metaphases from foetal tissues from the foetal offspring of X-irradiated female rats (5 Gy, acute dose) were analysed. The study's results showed that there is radiation-induced genomic instability: the number of aberrant metaphases and the breaks per total metaphases studied increased and were found to be statistically significant (p {<=} 0.05), with regard to the control group. In order to identify how this trans-generational, radiation-induced chromosomal instability could influence the chromosomal behaviour of the offspring of irradiated rat females in front of a chemical agent (aphidicolin), a total of 2481 metaphases were studied. The observed results showed that there is an enhancement of the action of the chemical agent: chromosomal breaks per aberrant metaphases show significant differences (p {<=} 0.05) in the X-ray- and aphidicolin-treated group as regards the aphidicolin-treated group. In conclusion, our findings indicate that there is trans-generational, radiation-induced chromosomal instability in the foetal

  11. Nicotinamide impairs entry into and exit from meiosis I in mouse oocytes.

    Directory of Open Access Journals (Sweden)

    Angelique Riepsamen

    Full Text Available Following exit from meiosis I, mammalian oocytes immediately enter meiosis II without an intervening interphase, accompanied by rapid reassembly of a bipolar spindle that maintains condensed chromosomes in a metaphase configuration (metaphase II arrest. Here we study the effect of nicotinamide (NAM, a non-competitive pan-sirtuin inhibitor, during meiotic maturation in mouse oocytes. Sirtuins are a family of seven NAD+-dependent deacetylases (Sirt1-7, which are involved in multiple cellular processes and are emerging as important regulators in oocytes and embryos. We found that NAM significantly delayed entry into meiosis I associated with delayed accumulation of the Cdk1 co-activator, cyclin B1. GVBD was also inhibited by the Sirt2-specific inhibitor, AGK2, and in a very similar pattern to NAM, supporting the notion that as in somatic cells, NAM inhibits sirtuins in oocytes. NAM did not affect subsequent spindle assembly, chromosome alignment or the timing of first polar body extrusion (PBE. Unexpectedly, however, in the majority of oocytes with a polar body, chromatin was decondensed and a nuclear structure was present. An identical phenotype was observed when flavopiridol was used to induce Cdk1 inactivation during late meiosis I prior to PBE, but not if Cdk1 was inactivated after PBE when metaphase II arrest was already established, altogether indicating that NAM impaired establishment rather than maintenance of metaphase II arrest. During meiosis I exit in NAM-treated medium, we found that cyclin B1 levels were lower and inhibitory Cdk1 phosphorylation was increased compared with controls. Although activation of the anaphase-promoting complex-Cdc20 (APC-Cdc20 occurred on-time in NAM-treated oocytes, Cdc20 levels were higher in very late meiosis I, pointing to exaggerated APC-Cdc20-mediated proteolysis as a reason for lower cyclin B1 levels. Collectively, therefore, our data indicate that by disrupting Cdk1 regulation, NAM impairs entry into

  12. Cytogenetic studies in 32 patients with myelodysplastic syndrome: insights to specific chromosomal abnormalities and prognosis.

    Science.gov (United States)

    Taniwaki, M; Horiike, S; Inazawa, J; Nishida, K; Misawa, S; Takino, T; Abe, T

    1987-06-01

    Cytogenetic studies were performed on a group of 32 patients with myelodysplastic syndrome (MDS). Five patients had refractory anemia (RA), four RA with ring sideroblasts (RARS), nine RA with excess blasts (RAEB), eight RAEB 'in transformation' (RAEB-T), three chronic myelomonocytic leukemia (CMML) and three secondary MDS (SMDS). Two patients in the SMDS group had been treated with alkylating agents for lung cancer and polycythemia vera, respectively. The other had been exposed to thorotrast. Chromosome analyses were performed with Q and G bandings on bone marrow cells incubated for 24 hr. A clonal chromosomal abnormality was observed in the marrow cells from 19 of the 32 patients (59%). Chromosomal abnormalities of nos. 5 and/or 7 were found in five patients, and were probably specific for RAEB-T and SMDS. Among the twelve patients with solely abnormal metaphases (AA), eight (67%) progressed to acute leukemia, a higher proportion than the three from the 13 patients (23%) with solely normal metaphases (NN) (P less than 0.05). One of the seven patients (14%) with both normal and abnormal metaphases (AN) developed acute leukemia (AA v. AN, P less than 0.03). In only two of the 12 patients who progressed to acute leukemia (17%), was complete remission achieved. The median survival time was only 4.0 months for patients with karyotype AA compared with 18.0 months for AN and 24.0 months for NN (AA v. AN, P less than 0.05, AA v. NN, P less than 0.05). The absence of cytogenetically normal cells indicated a poor prognosis with frequent progression to acute leukemia which is resistant to chemotherapy. Progression to acute leukemia depended not only on chromosomal abnormalities but also on morphological subtype classified according to French-American-British co-operative group criteria. Morphological findings and karyotype combined gave a good indication of the outcome for patients with MDS.

  13. PLD2 regulates microtubule stability and spindle migration in mouse oocytes during meiotic division

    Directory of Open Access Journals (Sweden)

    Xiaoyu Liu

    2017-05-01

    Full Text Available Phospholipase D2 (PLD2 is involved in cytoskeletal reorganization, cell migration, cell cycle progression, transcriptional control and vesicle trafficking. There is no evidence about PLD2 function in oocytes during meiosis. Herein, we analyzed PLD2 expression and its relationship with spindle formation and positioning in mouse oocyte meiosis. High protein level of PLD2 was revealed in oocytes by Western blot, which remained consistently stable from prophase I with intact germinal vesicle (GV up to metaphase II (MII stage. Immunofluorescence showed that PLD2 appeared and gathered around the condensed chromosomesafter germinal vesicle breakdown (GVBD, and co-localized with spindle from pro-metaphase I (pro-MI to metaphase I (MI and at MII stage. During anaphase I (Ana I to telophase I (Tel I transition, PLD2 was concentrated in the spindle polar area but absent from the midbody. In oocytes incubated with NFOT, an allosteric and catalytic inhibitor to PLD2, the spindle was enlarged and center-positioned, microtubules were resistant to cold-induced depolymerization and, additionally, the meiotic progression was arrested at MI stage. However, spindle migration could not be totally prevented by PLD2 catalytic specific inhibitors, FIPI and 1-butanol, implying at least partially, that PLD2 effect on spindle migration needs non-catalytic domain participation. NFOT-induced defects also resulted in actin-related molecules’ distribution alteration, such as RhoA, phosphatidylinosital 4, 5- biphosphate (PIP2, phosphorylated Colifin and, consequently, unordered F-actin dynamics. Taken together, these data indicate PLD2 is required for the regulation of microtubule dynamics and spindle migration toward the cortex in mammalian oocytes during meiotic progression.

  14. Complex Karyotype is a Stronger Predictor than Del(17p) for Inferior Outcome in Relapsed or Refractory CLL Patients Treated with Ibrutinib-Based Regimens

    Science.gov (United States)

    Thompson, Philip A.; O’Brien, Susan M.; Wierda, William G.; Ferrajoli, Alessandra; Stingo, Francesco; Smith, Susan C.; Burger, Jan A.; Estrov, Zeev; Jain, Nitin; Kantarjian, Hagop M.; Keating, Michael J.

    2016-01-01

    Background Ibrutinib is active in patients with relapsed/refractory (R/R) CLL. In patients treated with ibrutinib for R/R CLL, del(17p) identified by interphase fluorescence in situ hybridization (FISH) is associated with inferior progression-free survival, despite equivalent initial response rates. Del(17p) is frequently associated with complex metaphase karyotype (CKT); the prognostic significance of CKT in ibrutinib-treated patients has not been reported. Methods We reviewed 88 patients treated for R/R CLL at MD Anderson Cancer Center with investigational ibrutinib-based regimens from 2010–2013. Pre-treatment FISH and Lipopolysaccharide-stimulated metaphase cytogenetic analysis were performed on bone marrow. Results Adequate pre-treatment metaphase karyotype was available for 56/88 patients. Karyotype was complex in 21 of 56 cases; 17 of the 21 had del(17p) by FISH. Overall response rate, including partial remission with persistent lymphocytosis, was 94% with 17% complete responses. In multivariable analysis (MVA), only CKT was significantly associated with event-free survival (EFS) [HR 6.6 (1.7–25.6), p=0.006]. Fludarabine-refractory CLL [HR 6.9 (1.8–27.1), p=0.005] and CKT [HR 5.9 (1.6–22.2), p=0.008] were independently associated with inferior overall survival (OS) in MVA. Del(17p) by FISH was not significantly associated with EFS or OS in MVA. Conclusions CKT is a powerful predictor of outcome in ibrutinib-treated patients with R/R CLL and may be a stronger predictor of biological behavior than del(17p) by FISH. Given their relatively poor outcomes, patients with CKT are ideal candidates for studies of consolidative treatment strategies or novel treatment combinations. PMID:26193999

  15. Does Bariatric Surgery Improve Assisted Reproductive Technology Outcomes in Obese Infertile Women?

    Science.gov (United States)

    Milone, Marco; Sosa Fernandez, Loredana M; Sosa Fernandez, Laura V; Manigrasso, Michele; Elmore, Ugo; De Palma, Giovanni D; Musella, Mario; Milone, Francesco

    2017-08-01

    Little is known about the impact of bariatric surgery on obese infertile women seeking an assisted reproductive technology. All obese women with a history of assisted reproductive technology (ART) failure that underwent ART treatment cycles both prior to and following bariatric surgery were included. ART outcomes were compared evaluating the duration and dose of gonadotrophins used; the measurement of day 3 FSH; the anti-mullerian hormone dosage; the number of follicles >15 mm; the number of retrieved and fertilized oocytes; the number of metaphase II, metaphase I and germinal vesicle oocytes; the number of embryos obtained; the number of top-quality oocytes and embryos; the number of transferred embryo; the pregnancy rate and the live birth rate. Forty women were included. The total number of gonadotropin units required and in the length of stimulation following bariatric surgery decreased (p = .001), with an increase of the number of follicles ≥15 mm (p = .005), of retrieved oocytes (p = .004), of top-quality oocytes (p = .001) and metaphase II oocytes (p = .008). More oocytes were fertilized (4.2 ± 1.7 vs 5.3 ± 2.4; p = .02). After surgery, we have registered also a better number of top-quality embryos (0.5 ± 0.6 vs 1.1 ± 0.9; p = .003). Pregnancy rate following the bariatric surgery increased to 15/40 (37.5%) (p women seeking an ART treatment.

  16. Induction of Genomic Instability In Vivo by Low Doses of 137Cs gamma rays

    International Nuclear Information System (INIS)

    Rithidech, Kanokporn; Simon, Sanford R.; Whorton, Elbert B.

    2006-01-01

    The overall goal of this project is to determine if low doses (below or equal to the level traditionally requiring human radiation protection, i.e. less than or equal to 10 cGy) of low LET radiation can induce genomic instability. The magnitude of genomic instability was measured as delayed chromosome instability in bone marrow cells of exposed mice with different levels of endogenous DNA-dependent protein kinase catalytic subunit (DNA-PKcs) activity, i.e. high (C57BL/6J mice), intermediate (BALB/cJ mice), and extremely low (Scid mice). In addition, at early time points (1 and 4 hrs) following irradiation, levels of activation of nuclear factor-kappa B (NF-κB), a transcription factor known to be involved in regulating the expression of genes responsible for cell protection following stimuli, were measured in these cells. Bone marrow cells were collected at different times following irradiation, i.e. 1 hr, 4 hrs, 1 month, and 6 months. A total of five mice per dose per strain were sacrificed at each time point for sample collection. As a result, a total of 80 mice from each strain were used. The frequency and the type of metaphase chromosome aberrations in bone marrow cells collected from exposed mice at different times following irradiation were used as markers for radiation-induced genomic instability. A three-color fluorescence in situ hybridization (FISH) protocol for mouse chromosomes 1, 2, and 3 was used for the analysis of delayed stable chromosomal aberrations in metaphase cells. All other visible chromatid-type aberrations and gross structural abnormalities involving non-painted chromosomes were also evaluated on the same metaphase cells used for scoring the stable chromosomal aberrations of painted chromosomes. Levels of nuclear factor-kappa B (NF-κB) activation were also determined in cells at 1 and 4 hrs following irradiation (indicative of early responses)

  17. Hexavalent chromium is cytotoxic and genotoxic to hawksbill sea turtle cells

    International Nuclear Information System (INIS)

    Wise, Sandra S.; Xie, Hong; Fukuda, Tomokazu; Douglas Thompson, W.

    2014-01-01

    Sea turtles are a charismatic and ancient ocean species and can serve as key indicators for ocean ecosystems, including coral reefs and sea grass beds as well as coastal beaches. Genotoxicity studies in the species are absent, limiting our understanding of the impact of environmental toxicants on sea turtles. Hexavalent chromium (Cr(VI)) is a ubiquitous environmental problem worldwide, and recent studies show it is a global marine pollutant of concern. Thus, we evaluated the cytotoxicity and genotoxicity of soluble and particulate Cr(VI) in hawksbill sea turtle cells. Particulate Cr(VI) was both cytotoxic and genotoxic to sea turtle cells. Concentrations of 0.1, 0.5, 1, and 5 μg/cm 2 lead chromate induced 108, 79, 54, and 7% relative survival, respectively. Additionally, concentrations of 0, 0.1, 0.5, 1, and 5 μg/cm 2 lead chromate induced damage in 4, 10, 15, 26, and 36% of cells and caused 4, 11, 17, 30, and 56 chromosome aberrations in 100 metaphases, respectively. For soluble Cr, concentrations of 0.25, 0.5, 1, 2.5, and 5 μM sodium chromate induced 84, 69, 46, 25, and 3% relative survival, respectively. Sodium chromate induced 3, 9, 9, 14, 21, and 29% of metaphases with damage, and caused 3, 10, 10, 16, 26, and 39 damaged chromosomes in 100 metaphases at concentrations of 0, 0.25, 0.5, 1, 2.5, and 5 μM sodium chromate, respectively. These data suggest that Cr(VI) may be a concern for hawksbill sea turtles and sea turtles in general. - Highlights: • Particulate Cr(VI) is cytotoxic and clastogenic to hawksbill sea turtle cells. • Soluble Cr(VI) is cytotoxic and clastogenic to hawksbill sea turtle cells. • Cr(VI) may be a risk factor for hawksbill sea turtle health

  18. Drug and radiation sensitivity measurements of successful primary monolayer culturing of human tumor cells using cell-adhesive matrix and supplemented medium

    International Nuclear Information System (INIS)

    Baker, F.L.; Spitzer, G.; Ajani, J.A.

    1986-01-01

    The limitations of the agar suspension culture method for primary culturing of human tumor cells prompted development of a monolayer system optimized for cell adhesion and growth. This method grew 83% of fresh human tumor cell biopsy specimens, cultured and not contaminated, from a heterogeneous group of 396 tumors including lung cancer (93 of 114, 82%); melanoma (54 of 72, 75%); sarcoma (46 of 59, 78%); breast cancer (35 of 39, 90%); ovarian cancer (16 of 21, 76%); and a miscellaneous group consisting of gastrointestinal, genitourinary, mesothelioma, and unknown primaries (78 of 91, 86%). Cell growth was characterized morphologically with Papanicolaoustained coverslip cultures and cytogenetically with Giemsastained metaphase spreads. Morphological features such as nuclear pleomorphism, chromatin condensation, basophilic cytoplasm, and melanin pigmentation were routinely seen. Aneuploid metaphases were seen in 90% of evaluable cultures, with 15 of 28 showing 70% or more aneuploid metaphases. Colony-forming efficiency ranged between 0.01 and 1% of viable tumor cells, with a median efficiency of 0.2%. This culture system uses a low inoculum of 25,000 viable cells per well which permitted chemosensitivity testing of nine drugs at four doses in duplicate from 2.2 X 10(6) viable tumor cells and radiation sensitivity testing at five doses in quadruplicate from 0.6 X 10(6) cells. Cultures were analyzed for survival by computerized image analysis of crystal violet-stained cells. Drug sensitivity studies showed variability in sensitivity and in survival curve shape with exponential cell killing for cisplatin, Adriamycin, and etoposide, and shouldered survival curves for 5-fluorouracil frequently seen. Radiation sensitivity studies also showed variability in both sensitivity and survival curve shape. Many cultures showed exponential cell killing, although others had shouldered survival curves

  19. Shortening of culture time in conventional cytogenetic dosimetry

    International Nuclear Information System (INIS)

    Lamadrid, Ana I.; Gonzalez, Jorge E.; Romero, Ivonne; Garcia, Omar; Roy, Laurence

    2008-01-01

    Conventional cytogenetic dosimetry based on chromosome aberration in metaphases is a 'gold standard' of bio-dosimetry techniques for radiation dose assessment. This method is laborious and time consuming, the culturing process requires about 48 hours to obtain a satisfactory number of lymphocytes in mitosis. The current approach to reduce the dose estimation time by cytogenetic dosimetry is the preliminary estimation of dose counting only 50 metaphases. Another possibility is to reduce the culture time. The possibility of reduce the culture time under 48 hours adding Calyculin A has been suggested recently. In the present study we tested shorter times using Calyculin A and considering the G2/M-PCC index as culture quality indicator. Peripheral blood from healthy individuals was irradiated and then maintained at 37 C degrees for 2 hours allowing to act the cellular reparation mechanisms, lymphocytes were culture in RPMI 1640 supplemented with foetal calf serum and phytohemagglutinin. Colcemid was added 24 hours after cultures started and Calyculin A was added for the last hour. The cells were collected by centrifugation between 30 to 48 hours. The cells were treated with a hypotonic solution and the fixed cells dropped onto slides. The slides were stained with Giemsa. The incidence of metaphases with chromosomes well defined was scored. Two operators participated to the scoring according the same criteria. The results were analyzed to comparing the G2/M-PCC index relatives to achieve the shortest culture duration. The culture time reduction to 40 hours gives enough G2/M-PCC cells for dose estimation analysis. Lower culture times produced very low G2/M-PCC index. (author)

  20. Hexavalent Chromium Is Cytotoxic and Genotoxic to Hawksbill Sea Turtle Cells

    Science.gov (United States)

    Wise, Sandra S.; Xie, Hong; Fukuda, Tomokazu; Thompson, W. Douglas; Wise, John Pierce

    2014-01-01

    Sea turtles are a charismatic and ancient ocean species and can serve as key indicators for ocean ecosystems, including coral reefs and sea grass beds as well as coastal beaches. Genotoxicity studies in the species are absent, limiting our understanding of the impact of environmental toxicants on sea turtles. Hexavalent chromium (Cr(VI)) is a ubiquitous environmental problem worldwide, and recent studies show it is a global marine pollutant of concern. Thus, we evaluated the cytotoxicity and genotoxicity of soluble and particulate Cr(VI) in hawksbill sea turtle cells. Particulate Cr(VI) was both cytotoxic and genotoxic to sea turtle cells. Concentrations of 0.1, 0.5, 1, and 5 μg/cm2 lead chromate induced 108, 79, 54, and 7 percent relative survival, respectively. Additionally, concentrations of 0, 0.1, 0.5, 1, and 5 μg/cm2 lead chromate induced damage in 4, 10, 15, 26, and 36 percent of cells and caused 4, 11, 17, 30, and 56 chromosome aberrations in 100 metaphases, respectively. For soluble Cr, concentrations of 0.25, 0.5, 1, 2.5, and 5 μM sodium chromate induced 84, 69, 46, 25, and 3 percent relative survival, respectively. Sodium chromate induced 3, 9, 9, 14, 21, and 29 percent of metaphases with damage, and caused 3, 10, 10, 16, 26, and 39 damaged chromosomes in 100 metaphases at concentrations of 0, 0.25, 0.5, 1, 2.5, and 5 μM sodium chromate, respectively. These data suggest that Cr(VI) may be a concern for hawksbill sea turtles and sea turtles in general. PMID:24952338

  1. Dual trigger with gonadotropin-releasing hormone agonist and recombinant human chorionic gonadotropin improves in vitro fertilization outcome in gonadotropin-releasing hormone antagonist cycles.

    Science.gov (United States)

    Seval, Mehmet Murat; Özmen, Batuhan; Atabekoğlu, Cem; Şükür, Yavuz Emre; Şimşir, Coşkun; Kan, Özgur; Sönmezer, Murat

    2016-09-01

    The aim of this study was to evaluate whether dual trigger with leuprolide acetate plus recombinant human chorionic gonadotropin (hCG) improves in vitro fertilization outcome in gonadotropin-releasing hormone antagonist cycles. A total of 156 patients diagnosed with mild male factor, unexplained or tubal factor infertility were enrolled in the study. All subjects were allocated into one of two groups: the dual trigger group (leuprolide acetate 500 μg + recombinant hCG 250 μg) and the standard group (recombinant hCG 250 μg) according to the selected trigger method. Oocyte trigger was performed when at least three follicles >17 mm were observed. Pregnancy rate, number of collected oocytes, number of metaphase II oocytes, number of grade-A embryos, cycle cancellation rate, and ovarian hyperstimulation syndrome rate were the main outcome measures for the study. The mean number of grade-A embryos (1.6 ± 1.5 vs 1.1 ± 1.4, P = 0.01) and of metaphase II oocytes (7.9 ± 4.6 vs 6.3 ± 5.8, P = 0.02) was significantly higher in the dual-trigger group. Pregnancy rate was significantly higher in the dual-trigger group than in the standard group (54.8 vs 37.5%, P = 0.006). Two cases of mild ovarian hyperstimulation syndrome were observed in each group. This novel and more physiological trigger approach using 500 μg leuprolide acetate plus 250 μg recombinant hCG may lead to an increase in the number of metaphase II oocytes, grade-A embryos, and may improve pregnancy rates. © 2016 Japan Society of Obstetrics and Gynecology.

  2. PLK1 regulates spindle formation kinetics and APC/C activation in mouse zygote.

    Science.gov (United States)

    Baran, Vladimir; Brzakova, Adela; Rehak, Pavol; Kovarikova, Veronika; Solc, Petr

    2016-06-01

    Polo-like kinase 1 (PLK1) is involved in essential events of cell cycle including mitosis in which it participates in centrosomal microtubule nucleation, spindle bipolarity establishment and cytokinesis. Although PLK1 function has been studied in cycling cancer cells, only limited data are known about its role in the first mitosis of mammalian zygotes. During the 1-cell stage of mouse embryo development, the acentriolar spindle is formed and the shift from acentriolar to centrosomal spindle formation progresses gradually throughout the preimplantation stage, thus providing a unique possibility to study acentriolar spindle formation. We have shown previously that PLK1 activity is not essential for entry into first mitosis, but is required for correct spindle formation and anaphase onset in 1-cell mouse embryos. In the present study, we extend this knowledge by employing quantitative confocal live cell imaging to determine spindle formation kinetics in the absence of PLK1 activity and answer the question whether metaphase arrest at PLK1-inhibited embryos is associated with low anaphase-promoting complex/cyclosome (APC/C) activity and consequently high securin level. We have shown that inhibition of PLK1 activity induces a delay in onset of acentriolar spindle formation during first mitosis. Although these PLK1-inhibited 1-cell embryos were finally able to form a bipolar spindle, not all chromosomes were aligned at the metaphase equator. PLK1-inhibited embryos were arrested in metaphase without any sign of APC/C activation with high securin levels. Our results document that PLK1 controls the onset of spindle assembly and spindle formation, and is essential for APC/C activation before anaphase onset in mouse zygotes.

  3. Prenatal diagnosis and molecular cytogenetic characterization of a de novo 4.858-Mb microdeletion in 15q14 associated with ACTC1 and MEIS2 haploinsufficiency and tetralogy of Fallot.

    Science.gov (United States)

    Chen, Chih-Ping; Chen, Chen-Yu; Chern, Schu-Rern; Wu, Peih-Shan; Chen, Yen-Ni; Chen, Shin-Wen; Chen, Li-Feng; Yang, Chien-Wen; Wang, Wayseen

    2016-04-01

    To present prenatal diagnosis and molecular cytogenetic characterization of a de novo 15q14 microdeletion associated with tetralogy of Fallot (TOF). This was the first pregnancy of a 31-year-old primigravid woman. The pregnancy was uneventful until 23 weeks of gestation when TOF was first noted. The woman underwent amniocentesis at 23 weeks of gestation. Conventional cytogenetic analysis was performed using cultured amniocytes and parental bloods. Array comparative genomic hybridization (aCGH) was performed on uncultured amniocytes and parental bloods. Metaphase fluorescence in situ hybridization (FISH) was performed on cultured amniocytes. Quantitative fluorescent-polymerase chain reaction (QF-PCR) analysis was performed on placental tissue and parental bloods. Conventional cytogenetics on cultured amniocytes revealed a karyotype of 46,XY, aCGH on uncultured amniocytes revealed a de novo 4.858-Mb microdeletion in 15q14 encompassing ACTC1 and MEIS2, and metaphase FISH analysis on cultured amniocytes confirmed a 15q14 microdeletion. Postnatal phenotype included facial dysmorphism. QF-PCR assays detected a paternal origin of the 15q14 microdeletion in the fetus. Fetuses with 15q14 microdeletion may present TOF on the second-trimester ultrasound. aCGH and metaphase FISH are useful for rapid prenatal diagnosis of 15q14 microdeletion associated with TOF. A prenatal diagnosis of TOF should include a differential diagnosis of 15q14 microdeletion in addition to 22q11.2 deletion syndrome and other microdeletion syndromes. Copyright © 2016. Published by Elsevier B.V.

  4. Transforming growth factor beta-1 decreases the yield of the second meiotic division of rat pachytene spermatocytes in vitro

    Directory of Open Access Journals (Sweden)

    Sabido Odile

    2005-06-01

    Full Text Available Abstract Background TGF beta and its receptors are present in both germ cells and somatic cells of the male gonad. However, knock-out strategies for studying spermatogenesis regulation by TGF beta have been disappointing since TGF beta-or TGF beta receptor-null mice do not survive longer than a few weeks. Methods In the present study, we addressed the role of TGF beta-1 on the completion of meiosis by rat pachytene spermatocytes (PS cocultured with Sertoli cells. Identification and counting of meiotic cells were performed by cytology and cytometry. Results Under our culture conditions, some PS differentiated into round spermatids (RS. When TGF beta-1 was added to the culture medium, neither the number of PS or of secondary spermatocytes nor the half-life of RS was modified by the factor. By contrast, the number of RS and the amount of TP1 mRNA were lower in TGF beta-1-treated cultures than in control cultures. Very few metaphase I cells were ever observed both in control and TGF beta-1-treated wells. Higher numbers of metaphase II were present and their number was enhanced by TGF beta-1 treatment. A TGF beta-like bioactivity was detected in control culture media, the concentration of which increased with the time of culture. Conclusion These results indicate that TGF beta-1 did not change greatly, if any, the yield of the first meiotic division but likely enhanced a bottleneck at the level of metaphase II. Taken together, our results suggest strongly that TGF beta participates in an auto/paracrine pathway of regulation of the meiotic differentiation of rat spermatocytes.

  5. PARP inhibition causes premature loss of cohesion in cancer cells.

    Science.gov (United States)

    Kukolj, Eva; Kaufmann, Tanja; Dick, Amalie E; Zeillinger, Robert; Gerlich, Daniel W; Slade, Dea

    2017-11-28

    Poly(ADP-ribose) polymerases (PARPs) regulate various aspects of cellular function including mitotic progression. Although PARP inhibitors have been undergoing various clinical trials and the PARP1/2 inhibitor olaparib was approved as monotherapy for BRCA-mutated ovarian cancer, their mode of action in killing tumour cells is not fully understood. We investigated the effect of PARP inhibition on mitosis in cancerous (cervical, ovary, breast and osteosarcoma) and non-cancerous cells by live-cell imaging. The clinically relevant inhibitor olaparib induced strong perturbations in mitosis, including problems with chromosome alignment at the metaphase plate, anaphase delay, and premature loss of cohesion (cohesion fatigue) after a prolonged metaphase arrest, resulting in sister chromatid scattering. PARP1 and PARP2 depletion suppressed the phenotype while PARP2 overexpression enhanced it, suggesting that olaparib-bound PARP1 and PARP2 rather than the lack of catalytic activity causes this phenotype. Olaparib-induced mitotic chromatid scattering was observed in various cancer cell lines with increased protein levels of PARP1 and PARP2, but not in non-cancer or cancer cell lines that expressed lower levels of PARP1 or PARP2. Interestingly, the sister chromatid scattering phenotype occurred only when olaparib was added during the S-phase preceding mitosis, suggesting that PARP1 and PARP2 entrapment at replication forks impairs sister chromatid cohesion. Clinically relevant DNA-damaging agents that impair replication progression such as topoisomerase inhibitors and cisplatin were also found to induce sister chromatid scattering and metaphase plate alignment problems, suggesting that these mitotic phenotypes are a common outcome of replication perturbation.

  6. Cytogenetic effects of ionizing radiation in peripheral lymphocytes of ISS crew members

    Science.gov (United States)

    Johannes, Christian; Goedecke, Wolfgang; Antonopoulos, Alexandra; Obe, Günter; Horstmann, Markus

    High energy radiation is a major risk factor in manned space missions. Astronauts and cosmonauts are exposed to ionising radiations of cosmic and solar origin, while on the Earth's surface people are well protected by the atmosphere and a deflecting magnetic field. There are now data available describing the dose and the quality of ionising radiation on-board of the International Space Station (ISS). The effect of the increased radiation dose on mutation rates of ISS crew members are hard to predict. Therefore, direct measurements of mutation rates are required.The analysis of chromosomal aberrations in peripheral blood lymphocytes is a well established method to measure radiation-induced mutations. We present data of chromosome aberration analyses from lymphocyte metaphase spreads of ISS crew members participating in short term (10-14 days) or long term (6 months) missions. From each subject we received two blood samples. The first sample was drawn about 10 days before launch and a second sample was drawn within 3 days after return from their flights. From lymphocyte cultures metaphase plates were prepared on glass slides. Metaphases were Giemsa stained or hybridised using multicolour FISH probes. All types of chromosome changes were scored in pre-flight and post-flight blood samples and the mutation rates were compared. Results obtained in chromosomal studies on long-term flight crew members showed pronounced inter-individual differences in the response to cosmic radiation exposure. Overall significant elevations of typical radiation induced aberrations, i.e., dicentric chromosomes and reciprocal translocations have been observed in long-term crew members. Our data indicate no elevation of mutation rates due to short-term stays on-board the ISS.

  7. Rice ROOT ARCHITECTURE ASSOCIATED1 binds the proteasome subunit RPT4 and is degraded in a D-box and proteasome-dependent manner.

    Science.gov (United States)

    Han, Ye; Cao, Hong; Jiang, Jiafu; Xu, Yunyuan; Du, Jizhou; Wang, Xin; Yuan, Ming; Wang, Zhiyong; Xu, Zhihong; Chong, Kang

    2008-10-01

    Root growth is mainly determined by cell division and subsequent elongation in the root apical area. Components regulating cell division in root meristematic cells are largely unknown. Previous studies have identified rice (Oryza sativa) ROOT ARCHITECTURE ASSOCIATED1 (OsRAA1) as a regulator in root development. Yet, the function of OsRAA1 at the cellular and molecular levels is unclear. Here, we show that OsRAA1-overexpressed transgenic rice showed reduced primary root growth, increased numbers of cells in metaphase, and reduced numbers of cells in anaphase, which suggests that OsRAA1 is responsible for limiting root growth by inhibiting the onset of anaphase. The expression of OsRAA1 in fission yeast also induced metaphase arrest, which is consistent with the fact that OsRAA1 functions through a conserved mechanism of cell cycle regulation. Moreover, a colocalization assay has shown that OsRAA1 is expressed predominantly at spindles during cell division. Yeast two-hybrid and pull-down assays, as well as a bimolecular fluorescence complementation assay, all have revealed that OsRAA1 interacts with a rice homolog of REGULATORY PARTICLE TRIPLE-A ATPASE4, a component that is involved in the ubiquitin pathway. Treating transgenic rice with specific inhibitors of the 26S proteasome blocked the degradation of OsRAA1 and increased the number of cells in metaphase. Mutation of a putative ubiquitination-targeting D-box (RGSLDLISL) in OsRAA1 interrupted the destruction of OsRAA1 in transgenic yeast. These results suggest that ubiquitination and proteasomic proteolysis are involved in OsRAA1 degradation, which is essential for the onset of anaphase, and that OsRAA1 may modulate root development mediated by the ubiquitin-proteasome pathway as a novel regulatory factor of the cell cycle.

  8. Expedited Radiation Biodosimetry by Automated Dicentric Chromosome Identification (ADCI) and Dose Estimation.

    Science.gov (United States)

    Shirley, Ben; Li, Yanxin; Knoll, Joan H M; Rogan, Peter K

    2017-09-04

    Biological radiation dose can be estimated from dicentric chromosome frequencies in metaphase cells. Performing these cytogenetic dicentric chromosome assays is traditionally a manual, labor-intensive process not well suited to handle the volume of samples which may require examination in the wake of a mass casualty event. Automated Dicentric Chromosome Identifier and Dose Estimator (ADCI) software automates this process by examining sets of metaphase images using machine learning-based image processing techniques. The software selects appropriate images for analysis by removing unsuitable images, classifies each object as either a centromere-containing chromosome or non-chromosome, further distinguishes chromosomes as monocentric chromosomes (MCs) or dicentric chromosomes (DCs), determines DC frequency within a sample, and estimates biological radiation dose by comparing sample DC frequency with calibration curves computed using calibration samples. This protocol describes the usage of ADCI software. Typically, both calibration (known dose) and test (unknown dose) sets of metaphase images are imported to perform accurate dose estimation. Optimal images for analysis can be found automatically using preset image filters or can also be filtered through manual inspection. The software processes images within each sample and DC frequencies are computed at different levels of stringency for calling DCs, using a machine learning approach. Linear-quadratic calibration curves are generated based on DC frequencies in calibration samples exposed to known physical doses. Doses of test samples exposed to uncertain radiation levels are estimated from their DC frequencies using these calibration curves. Reports can be generated upon request and provide summary of results of one or more samples, of one or more calibration curves, or of dose estimation.

  9. Hexavalent chromium is cytotoxic and genotoxic to hawksbill sea turtle cells

    Energy Technology Data Exchange (ETDEWEB)

    Wise, Sandra S., E-mail: sandra.wise@maine.edu [Wise Laboratory of Environmental and Genetic Toxicology, University of Southern Maine, Science Building, 96 Falmouth Street, Portland, ME 04103 (United States); Maine Center for Toxicology and Environmental Health, University of Southern Maine, Science Building, 96 Falmouth Street, Portland, ME 04103 (United States); Department of Applied Medical Science, University of Southern Maine, Science Building, 96 Falmouth Street, Portland, ME 04103 (United States); Xie, Hong, E-mail: hongxie@usm.maine.edu [Wise Laboratory of Environmental and Genetic Toxicology, University of Southern Maine, Science Building, 96 Falmouth Street, Portland, ME 04103 (United States); Maine Center for Toxicology and Environmental Health, University of Southern Maine, Science Building, 96 Falmouth Street, Portland, ME 04103 (United States); Department of Applied Medical Science, University of Southern Maine, Science Building, 96 Falmouth Street, Portland, ME 04103 (United States); Fukuda, Tomokazu, E-mail: tomofukuda009@gmail.com [Graduate School of Agricultural Sciences, Tohoku University, Laboratory of Animal Breeding and Genetics, Second Research Building, Rm 112, 1-1 Amamiyamachi, Aoba-ku, Sendai 981-8555 (Japan); Douglas Thompson, W., E-mail: dougt@usm.maine.edu [Maine Center for Toxicology and Environmental Health, University of Southern Maine, Science Building, 96 Falmouth Street, Portland, ME 04103 (United States); Department of Applied Medical Science, University of Southern Maine, Science Building, 96 Falmouth Street, Portland, ME 04103 (United States); and others

    2014-09-01

    Sea turtles are a charismatic and ancient ocean species and can serve as key indicators for ocean ecosystems, including coral reefs and sea grass beds as well as coastal beaches. Genotoxicity studies in the species are absent, limiting our understanding of the impact of environmental toxicants on sea turtles. Hexavalent chromium (Cr(VI)) is a ubiquitous environmental problem worldwide, and recent studies show it is a global marine pollutant of concern. Thus, we evaluated the cytotoxicity and genotoxicity of soluble and particulate Cr(VI) in hawksbill sea turtle cells. Particulate Cr(VI) was both cytotoxic and genotoxic to sea turtle cells. Concentrations of 0.1, 0.5, 1, and 5 μg/cm{sup 2} lead chromate induced 108, 79, 54, and 7% relative survival, respectively. Additionally, concentrations of 0, 0.1, 0.5, 1, and 5 μg/cm{sup 2} lead chromate induced damage in 4, 10, 15, 26, and 36% of cells and caused 4, 11, 17, 30, and 56 chromosome aberrations in 100 metaphases, respectively. For soluble Cr, concentrations of 0.25, 0.5, 1, 2.5, and 5 μM sodium chromate induced 84, 69, 46, 25, and 3% relative survival, respectively. Sodium chromate induced 3, 9, 9, 14, 21, and 29% of metaphases with damage, and caused 3, 10, 10, 16, 26, and 39 damaged chromosomes in 100 metaphases at concentrations of 0, 0.25, 0.5, 1, 2.5, and 5 μM sodium chromate, respectively. These data suggest that Cr(VI) may be a concern for hawksbill sea turtles and sea turtles in general. - Highlights: • Particulate Cr(VI) is cytotoxic and clastogenic to hawksbill sea turtle cells. • Soluble Cr(VI) is cytotoxic and clastogenic to hawksbill sea turtle cells. • Cr(VI) may be a risk factor for hawksbill sea turtle health.

  10. Bub3 is a spindle assembly checkpoint protein regulating chromosome segregation during mouse oocyte meiosis.

    Directory of Open Access Journals (Sweden)

    Mo Li

    Full Text Available In mitosis, the spindle assembly checkpoint (SAC prevents anaphase onset until all chromosomes have been attached to the spindle microtubules and aligned correctly at the equatorial metaphase plate. The major checkpoint proteins in mitosis consist of mitotic arrest-deficient (Mad1-3, budding uninhibited by benzimidazole (Bub1, Bub3, and monopolar spindle 1(Mps1. During meiosis, for the formation of a haploid gamete, two consecutive rounds of chromosome segregation occur with only one round of DNA replication. To pull homologous chromosomes to opposite spindle poles during meiosis I, both sister kinetochores of a homologue must face toward the same pole which is very different from mitosis and meiosis II. As a core member of checkpoint proteins, the individual role of Bub3 in mammalian oocyte meiosis is unclear. In this study, using overexpression and RNA interference (RNAi approaches, we analyzed the role of Bub3 in mouse oocyte meiosis. Our data showed that overexpressed Bub3 inhibited meiotic metaphase-anaphase transition by preventing homologous chromosome and sister chromatid segregations in meiosis I and II, respectively. Misaligned chromosomes, abnormal polar body and double polar bodies were observed in Bub3 knock-down oocytes, causing aneuploidy. Furthermore, through cold treatment combined with Bub3 overexpression, we found that overexpressed Bub3 affected the attachments of microtubules and kinetochores during metaphase-anaphase transition. We propose that as a member of SAC, Bub3 is required for regulation of both meiosis I and II, and is potentially involved in kinetochore-microtubule attachment in mammalian oocytes.

  11. H3 Thr3 phosphorylation is crucial for meiotic resumption and anaphase onset in oocyte meiosis.

    Science.gov (United States)

    Wang, Qian; Wei, Haojie; Du, Juan; Cao, Yan; Zhang, Nana; Liu, Xiaoyun; Liu, Xiaoyu; Chen, Dandan; Ma, Wei

    2016-01-01

    Haspin-catalyzed histone H3 threonine 3 (Thr3) phosphorylation facilitates chromosomal passenger complex (CPC) docking at centromeres, regulating indirectly chromosome behavior during somatic mitosis. It is not fully known about the expression and function of H3 with phosphorylated Thr3 (H3T3-P) during meiosis in oocytes. In this study, we investigated the expression and sub-cellular distribution of H3T3-P, as well as its function in mouse oocytes during meiotic division. Western blot analysis revealed that H3T3-P expression was only detected after germinal vesicle breakdown (GVBD), and gradually increased to peak level at metaphase I (MI), but sharply decreased at metaphase II (MII). Immunofluorescence showed H3T3-P was only brightly labeled on chromosomes after GVBD, with relatively high concentration across the whole chromosome axis from pro-metaphase I (pro-MI) to MI. Specially, H3T3-P distribution was exclusively limited to the local space between sister centromeres at MII stage. Haspin inhibitor, 5-iodotubercidin (5-ITu), dose- and time-dependently blocked H3T3-P expression in mouse oocytes. H3T3-P inhibition delayed the resumption of meiosis (GVBD) and chromatin condensation. Moreover, the loss of H3T3-P speeded up the meiotic transition to MII of pro-MI oocytes in spite of the presence of non-aligned chromosomes, even reversed MI-arrest induced with Nocodazole. The inhibition of H3T3-P expression distinguishably damaged MAD1 recruitment on centromeres, which indicates the spindle assembly checkpoint was impaired in function, logically explaining the premature onset of anaphase I. Therefore, Haspin-catalyzed histone H3 phosphorylation is essential for chromatin condensation and the following timely transition from meiosis I to meiosis II in mouse oocytes during meiotic division.

  12. SGO1 maintains bovine meiotic and mitotic centromeric cohesions of sister chromatids and directly affects embryo development.

    Directory of Open Access Journals (Sweden)

    Feng-Xia Yin

    Full Text Available Shugoshin (SGO is a critical factor that enforces cohesion from segregation of paired sister chromatids during mitosis and meiosis. It has been studied mainly in invertebrates. Knowledge of SGO(s in a mammalian system has only been reported in the mouse and Hela cells. In this study, the functions of SGO1 in bovine oocytes during meiotic maturation, early embryonic development and somatic cell mitosis were investigated. The results showed that SGO1 was expressed from germinal vesicle (GV to the metaphase II stage. SGO1 accumulated on condensed and scattered chromosomes from pre-metaphase I to metaphase II. The over-expression of SGO1 did not interfere with the process of homologous chromosome separation, although once separated they were unable to move to the opposing spindle poles. This often resulted in the formation of oocytes with 60 replicated chromosomes. Depletion of SGO1 in GV oocytes affected chromosomal separation resulting in abnormal chromosome alignment at a significantly higher proportion than in control oocytes. Knockdown of SGO1 expression significantly decreased the embryonic developmental rate and quality. To further confirm the function(s of SGO1 during mitosis, bovine embryonic fibroblast cells were transfected with SGO1 siRNAs. SGO1 depletion induced the premature dissociation of chromosomal cohesion at the centromere and along the chromosome arm giving rise to abnormal appearing mitotic patterns. The results of this study infer that SGO1 is involved in the centromeric cohesion of sister chromatids and chromosomal movement towards the spindle poles. Depletion of SGO1 causes arrestment of cell division in meiosis and mitosis.

  13. Prenatal diagnosis of the derivative chromosome 22 associated with cat eye syndrome by fluorescence in situ hybridization.

    Science.gov (United States)

    Reeser, S L; Donnenfeld, A E; Miller, R C; Sellinger, B S; Emanuel, B S; Driscoll, D A

    1994-11-01

    Cytogenetic studies of cultured amniocytes demonstrated a karyotype of 46,XX/47,XX, +mar. A bisatellited, dicentric, distamycin-DAPI negative, NOR-positive marker was present in 76 per cent of the metaphases examined. Similar markers have been associated with cat eye syndrome (CES). We report on the utilization of fluorescence in situ hybridization (FISH) with a 14/22 alpha-satellite probe and a chromosome 22-specific cosmid for locus D22S9 to determine the origin of the prenatally detected supernumerary marker chromosome. FISH studies demonstrated that the marker is a derivative of chromosome 22 and enabled us to provide the family with additional prognostic information.

  14. 46,XX Karyotype in a Male with Ambigious Genitalia: A Case Report

    Directory of Open Access Journals (Sweden)

    Mahmut Balkan

    2004-01-01

    Full Text Available 40 days old case who were diagnosed with ambigious genitalia were sentto laboratory of cytogenetics. Mother was 26 years old and have to alivechildren and case were the second child and applied to Child SurgeryDepartment due to continiously vomiting and uneasy conditions. Sexualdevelopment were as male on physical examinations. Cell culture wasapplied for chromosomal analysis. Slides were stained with GiemsaBanding Staining (GTG and 100 cells were totaly counted and karyotypingwere done with 15 metaphase. Chromosome with 46,XX karyotype. Casewere taken under consideration of congenital adrenel hyperplasia afterevalution of karyotype. Case were discussed according to by information ofpresents literatures.

  15. Karyotype study of the South American fruit fly, Anastrepha fraterculus (Wied.) in Argentina

    International Nuclear Information System (INIS)

    Lifschitz, E.; Manso, F.; Basso, A.

    1999-01-01

    The most frequent karyotype of Anastrepha fraterculus in Argentina is described here on the basis of mitotic metaphase morphology. It was named ''fraterculus Arg 1''. The diploid number is 2n=10+XX/XY and in males it comprises five homomorphic pairs and one heteomorphic pair, the latter being the sexual pair. Samples from different populations were cytologically analyzed, and ''fraterculus Arg 1'' is present in all of them at a high frequency (about 60%). A typical C band pattern of the X chromosome was found only in the Montecarlo (Misiones province) population. (author)

  16. Identification of novel translocation between short arm of chromosome 4 and long arm of chromosome 6 in an infertile man using Interphase Chromosome Profiling (ICP).

    Science.gov (United States)

    Kaul, S; Kaur, H; Vats, S K S; Chawla, J; Jindal, R; Khetarpal, P

    2018-02-07

    Conventional cytogenetics has always been a favourite to detect chromosomal aberrations. Carriers of chromosomal translocation are often phenotypically normal but are infertile. Couples are often advised to go for karyotyping, but culture failure or improper metaphase spread with poor banding often makes the analysis difficult. We report here a novel translocation between short arm of chromosome 4 and long arm of chromosome 6 in an infertile man using an advanced molecular cytogenetic technique of Interphase Chromosome Profiling (ICP). © 2018 Blackwell Verlag GmbH.

  17. A parallel approach for subwavelength molecular surgery using gene-specific positioned metal nanoparticles as laser light antennas.

    Science.gov (United States)

    Csaki, Andrea; Garwe, Frank; Steinbrück, Andrea; Maubach, Gunter; Festag, Grit; Weise, Anja; Riemann, Iris; König, Karsten; Fritzsche, Wolfgang

    2007-02-01

    An optical technique for the parallel manipulation of nanoscale structures with molecular resolution is presented. Bioconjugated metal nanoparticles are thereby positioned at the location of interest, such as, e.g., certain DNA sequences along metaphase chromosomes, prior to pulsed laser light irradiation of the whole sample. The nanoparticles are designed to absorb the introduced energy highly efficiently, in that way acting as nanoantenna. As result of the interaction, structural changes of the sample with subwavelength dimensions and nanoscale precision are observed at the location of the particles. The process leading to the nanolocalized destruction is caused by particle ablation as well as thermal damage of the surrounding material.

  18. THE CYTOGENETIC EFFECTS WITH LEAD NITROGENOUS TREATMENT AT HELIANTHUS ANNUUS L. FAVORIT AND PR64A83 CULTIVARS

    Directory of Open Access Journals (Sweden)

    Felicia Vlad-Rusen

    2007-08-01

    Full Text Available The analysis of some cytogenetic parameters shows that the nitrogenous lead application on Favorit and PR64A83 cultivars of sunflower exercises a light mutagenic effect on the cells of the root apex. At same time an intensification of the frequency of the ana-telophases with aberration takes place, especially of those with bridges, with lagging chromosomes, but also of those with fragments, demonstrating the perturbatory action that this substance exercises on the division axle. The apparition of a great number of metaphases with displayed chromosomes proves the perturbation of well function to division axle, effect signalized especially at the Favorit cultivar.

  19. Genetic lesions induced by chemicals in spermatozoa and spermatids of mice are repaired in the egg.

    Science.gov (United States)

    Generoso, W M; Cain, K T; Krishna, M; Huff, S W

    1979-01-01

    Conclusive proof that the mouse egg is capable of carrying out repair of genetic lesions present in the male genome was obtained through dominant-lethal studies of chemically treated spermatozoa and spermatids and through cytological analysis of first-cleavage metaphases. The maximum difference in repair capability between stocks of females, found for isopropyl methanesulfonate treatment, was large; considerably smaller differences were found for ethyl methanesulfonate, triethylenemelamine, and benzo[a]pyrene treatments; and no difference was found for x-ray treatment.

  20. Differential answer the two main types radioinduced chromosomal aberrations in front of two inhibitors the reparative synthesis

    International Nuclear Information System (INIS)

    Lopez, I.; Pincheira, J.

    1998-01-01

    One studies the effect two inhibitors the reparative DNA synthesis: afidicolina (Apc) and dideoxitimidina (ddThd) in the frequency dicentric and translocases in lymphocytes irradiated with 3 Gy gives rays X. The estimate aberrations one carries out in metaphases banded G, obtained by microcultivo outlying blood a giver, irradiated in presence and absence gives different concentrations Apc and ddThd. The obtained results could indicate that the processes give erroneous repair that lead to the formation dicentric and translocases, that dicentric no and translocases could involve to different polimerasas

  1. Brodie’s Abscess in a Patient Presenting with Sickle Cell Vasoocclusive Crisis

    Directory of Open Access Journals (Sweden)

    Onyekachi Henry Ogbonna

    2015-01-01

    Full Text Available First described by Sir Nicholas Brodie in 1832, Brodie’s abscess is a localized subacute or chronic infection of the bone, typically seen in the metaphases of long bones in children and adolescents. The diagnosis can prove to be enigmatic due to absence of clinical signs and symptoms of systemic disease. We report a very interesting case of Brodie’s abscess masquerading as sickle cell vasoocclusive crisis in a 20-year-old female with sickle cell disease and review the literature.

  2. Biological radiation dose estimation by chromosomal aberrations analysis in human peripheral blood (dose- effect curve)

    International Nuclear Information System (INIS)

    Al Achkar, W.

    2002-01-01

    In order to draw a dose-effect curve, blood from eight healthy people were studied. Samples were irradiated in tubes with 0.15-2.5 gray of gamma ray.Irradiated and control samples were incubated for cell cultures. Chromosomal aberrations from 67888 metaphases were scored. Curves from the total number of dicentrics, dicentrics+ rings and total numbers of breaks were drawn. The yield of chromosome aberrations is related to the dose used. These curves give a quick useful estimation of the accidentally radiation exposure. (author)

  3. Detection of Alien Oryza punctata Kotschy Chromosomes in Rice, Oryza sativa L., by Genomic in situ Hybridization

    OpenAIRE

    Yasui, Hideshi; Nonomura, Ken-ichi; Iwata, Nobuo; 安井, 秀; 野々村, 賢一; 岩田, 伸夫

    1997-01-01

    Genomic in situ hybridization (GIS H) using total Oryza punctata Kotschy genomic DNA as a probe was applied to detect alien chromosomes transferred from O. punctata (W1514: 2n=2x=24: BB) to O. sativa Japonica cultivar, Nipponbare (2n=2x=24: AA). Only 12 chromosomes in the interspecific hybrids (2n=3x=36: AAB) between autotetraploid of O. sativa cultivar Nipponbare and a diploid strain of O. punctata (W1514) showed intense staining by FITC in mitotic metaphase spreads. Only one homologous pair...

  4. Comparative proteomics of mitosis and meiosis in Saccharomyces cerevisiae.

    Science.gov (United States)

    Kumar, Ravinder; Dhali, Snigdha; Srikanth, Rapole; Ghosh, Santanu Kumar; Srivastava, Sanjeeva

    2014-09-23

    Precise and timely segregation of genetic material and conservation of ploidy are the two foremost requirements for survival of a eukaryotic organism. Two highly regulated cell division processes, namely mitosis and meiosis are central to achieve this objective. The modes of chromosome segregation are distinct in these two processes that generate progeny cells of equal ploidy and half the ploidy in mitosis and meiosis, respectively. Additionally, the nutritional requirement and intracellular processing of biological cue also differ in these two processes. From this, it can be envisaged that proteome of mitotic and meiotic cells will differ significantly. Therefore, identification of proteins that differ in their level of expression between mitosis and meiosis would further reveal the mechanistic detail of these processes. In the present study, we have investigated the protein expression profile of mitosis and meiosis by comparing proteome of budding yeast cultures arrested at mitotic metaphase and metaphase-I of meiosis using proteomic approach. Approximately 1000 and 2000 protein spots were visualized on 2-DE and 2D-DIGE gels respectively, out of which 14 protein spots were significant in 2-DE and 22 in 2D-DIGE (pmitosis, an up-regulation of actin cytoskeleton and its negative regulator occurs in meiosis. Mitosis and meiosis are two different types of cell division cycles with entirely different outcomes with definite biological implication for almost all eukaryotic species. In this work, we investigated, for the first time, the differential proteomic profile of Saccharomyces cerevisiae culture arrested at mitotic metaphase (M) and metaphase-I (MI) of meiosis using 2-DE and 2D-DIGE. Our findings of up-regulation of actin and its negative regulator cofilin during meiosis suggest that the rate of actin cytoskeleton turnover is more in meiosis and actin cytoskeleton may play more crucial role during meiosis compared to mitosis. Present study also suggests that actin

  5. Assignment of a Polycomb-like chromobox gene (CBX2) to human chromosome 17q25

    Energy Technology Data Exchange (ETDEWEB)

    Gecz, J.; Gaunt, S.J.; Passage, E. [INSERM, Marseille (France)] [and others

    1995-03-01

    A human clone corresponding to the homolgue of the murine Polycomb-like gene M33 has been used to map this gene (CBX2) to human chromosomes. Both somatic cell hybrid panels and FISH on a metaphase chromosomes have been used. These techniques gave a consistent localization, at the tip of the long arm of chromosome 17 (17q25). This localization, as well as the the potential role of a mammalian Polycomb-like protein, suggests a potential involvement in two different pathologies: the campomelic syndrome, an inherited disorder, and neoplastic disorders linked to allele loss already described in this region. 15 refs., 4 figs.

  6. Serially cultured keratinocytes from human scalp hair follicles: a tool for cytogenetic studies.

    Science.gov (United States)

    Weterings, P J; Roelofs, H M; Jansen, B A; Vermorken, A J

    1983-01-01

    Keratinocytes originating from adult human hair follicles, the most convenient biopsy tissue, can be serially cultured using a combination of two techniques. Primary cultures are established using plucked scalp hair follicles and the bovine eye lens capsule as a growth substrate. Subsequently, cells from these cultures are serially cultivated in the presence of irradiated 3T3 cells as feeders. By this combination of techniques many keratinocytes can be generated from one single hair follicle. These cultures, appropriately treated with colchicine, can provide an adequate number of metaphases suitable for chromosome studies.

  7. Nanotomography of Cell Surfaces with Evanescent Fields

    Directory of Open Access Journals (Sweden)

    Michael Wagner

    2008-01-01

    Full Text Available The technique of variable-angle total internal reflection fluorescence microscopy (TIRFM and its application to nanotomography of cell surfaces are described. Present applications include (1 3D imaging of chromosomes in their metaphase to demonstrate axial resolution in the nanometre range, (2 measurements of cell-substrate topology, which upon cholesterol depletion shows some loosening of cell-substrate contacts, and (3 measurements of cell topology upon photodynamic therapy (PDT, which demonstrate cell swelling and maintenance of focal contacts. The potential of the method for in vitro diagnostics, but also some requirements and limitations are discussed.

  8. Birth after 12 hours of oocyte refrigeration.

    Science.gov (United States)

    Coban, Onder; Hacifazlioglu, Oguzhan; Ciray, H Nadir; Ulug, Ulun; Tekin, H Ibrahim; Bahceci, Mustafa

    2010-12-01

    To assess cycle outcome after oocyte refrigeration. Case report. Private IVF center. One couple in a donor oocyte program. Intracytoplasmic sperm injection and blastocyst culture after refrigeration of oocytes for 12 hours. Birth. Fourteen two-pronuclei zygotes from 17 metaphase II refrigerated oocytes resulted in transfer of two blastocysts at day 5 and cryopreservation of six excess embryos at day 6. The patient delivered one healthy male baby after 38 weeks' gestation. The successful outcome of oocyte refrigeration indicates that this protocol could be useful in circumstances in which a delay in obtaining spermatozoa arises. Copyright © 2010 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  9. Trisomy 19 and T(9;22 In a Patient with Acute Basophilic Leukemia

    Directory of Open Access Journals (Sweden)

    Alicia Rojas-Atencio

    2011-01-01

    Full Text Available We report a case of acute basophilic leukemia with two coexisting clonal abnormalities, t(9;22 and trisomy 19. The blast showed positive reaction with myeloperoxidase but negative reaction with chloroacetate esterase and acid phosphatase. Metachromatic features of the blast were observed with toluidine blue stain. Ultrastructure study showed the presence of azurophilic granules in basophils and blast mast cells. Conventional and molecular cytogenetic studies revealed, t(9;22 with BCR/ABL positive and trisomy 19 in all metaphase cells. To our knowledge, this paper here is the first to present acute basophilic leukemia with trisomy 19 and t(9;22.

  10. Effects of ionizing radiations on roses, 2

    International Nuclear Information System (INIS)

    Lata, P.

    1975-01-01

    A comparative study of the male meiosis in the rose cultivar ''Pink Parfait'' and its two, Deep and Light pink flowered mutants was conducted. ''Pink Parfait'' was found to be a segmental allotetraploid. The frequency of univalents and quadrivalents at metaphase I was higher in the mutants. Anaphase I revealed a higher incidence of lagging chromosomes in Deep pink mutant and an increased frequency of chromosome bridges in both the mutants. The deleterious effects of radiations were evident from the higher percentage of chromosomal aberrations and pollen sterility found in mutants as compared with the control. (auth.)

  11. Micromechanical study of protein-DNA interactions and chromosomes

    Science.gov (United States)

    Marko, John

    I will discuss micromechanics experiments that our group has used to analyze protein-DNA interactions and chromosome organization. In single-DNA experiments we have found that a feature of protein-DNA complexes is that their dissociation rates can depend strikingly on bulk solution concentrations of other proteins and DNA segments; I will describe experiments which demonstrate this effect, which can involve tens-fold changes in off-rates with submicromolar changes in solution concentrations. Second, I will discuss experiments aimed at analyzing large-scale human chromosome structure; we isolate metaphase chromosomes, which in their native form behave as remarkably elastic networks of chromatin. Exposure to DNA-cutting restriction enzymes completely eliminates this elasticity, indicating that there is not a mechanically contiguous protein ''scaffold'' from which the chromosome gains its stability. I will show results of siRNA experiments indicating that depletion of condensin proteins leads to destabilization of chromosome mechanics, indicating condensin's role as the major chromatin ''cross-linker'' in metaphase chromosomes. Finally I will discuss similar experiments on human G1 nuclei, where we use genetic and chemical modifications to separate the contributions of the nuclear lamina and chromatin to the mechanical stiffness of the nucleus as a whole. Supported by the NSF (DMR-1206868, MCB-1022117) and the NIH (GM105847, CA193419).

  12. Ergonomic audit of a specially engineered sonic powered toothbrush with unique sensing and control technologies, the Sonicare Flexcare, and the Oral-B Smart Series 5000.

    Science.gov (United States)

    Hunter, Gail; Burns, Laurie; Bone, Brian; Mintel, Thomas; Jimenez, Eduardo

    2012-01-01

    The presence of ergonomic features can impact the marketplace success of a new product. Metaphase Design Group, Inc., in partnership with the Colgate-Palmolive Company, conducted an ergonomic audit on three electric toothbrushes: a specially engineered sonic powered toothbrush with unique sensing and control technologies, the Sonicare FlexCare, and the Oral-B Smart Series 5000. The ergonomic audit was conducted by Metaphase Design Groups's ergonomic and usability experts. Two experts used the toothbrushes over a one-week period and assessed the performance of each brush against a set of ergonomic principles. The three toothbrushes have some solid ergonomic features. They each have adequate grip zones, provide grip security with elastomeric materials, and provide easy access to the on/off button. The most distinctive feature is the longitudinal shape of the handle of the specially engineered sonic powered toothbrush with unique sensing and control technologies. This handle angles downward at the top end and provides additional advantages through improved grip security and visibility. Yet all three toothbrushes have different opportunities for improvement. The Sonicare Flex Care toothbrush has a cluttered and complicated user interface that is difficult to read. The disadvantages of the Oral-B Smart Series 5000 toothbrush are related to its physical dimensions and audible feedback. The specially engineered sonic powered toothbrush with unique sensing afid control technologies is surprising to use with its changes in speeds, brush movements, and resulting changes in audible feedback.

  13. Replication Banding Patterns in Human Chromosomes Detected Using 5-ethynyl-2'-deoxyuridine Incorporation

    International Nuclear Information System (INIS)

    Hoshi, Osamu; Ushiki, Tatsuo

    2011-01-01

    A novel technique using the incorporation of 5-ethynyl-2'-deoxyuridine (EdU) into replicating DNA is described for the analysis of replicating banding patterns of human metaphase chromosomes. Human lymphocytes were synchronized with excess thymidine and treated with EdU during the late S phase of the cell cycle. The incorporated EdU was then detected in metaphase chromosomes using Alexa Fluor® 488 azides, through the 1,3-dipolar cycloaddition reaction of organic azides with the terminal acetylene group of EdU. Chromosomes with incorporated EdU showed a banding pattern similar to G-banding of normal human chromosomes. Imaging by atomic force microscopy (AFM) in liquid conditions showed that the structure of the chromosomes was well preserved even after EdU treatment. Comparison between fluorescence microscopy and AFM images of the same chromosome 1 indicated the presence of ridges and grooves in the chromatid arm, features that have been previously reported in relation to G-banding. These results suggest an intimate relationship between EdU-induced replication bands and G- or R-bands in human chromosomes. This technique is thus useful for analyzing the structure of chromosomes in relation to their banding patterns following DNA replication in the S phase

  14. Preparation of single rice chromosome for construction of a DNA library using a laser microbeam trap.

    Science.gov (United States)

    Liu, Xiaohui; Wang, Haowei; Li, Yinmei; Tang, Yesheng; Liu, Yilei; Hu, Xin; Jia, Peixin; Ying, Kai; Feng, Qi; Guan, Jianping; Jin, Chaoqing; Zhang, Lei; Lou, Liren; Zhou, Zhuan; Han, Bin

    2004-04-29

    We report the development of a laser micromanipulation system and its application in the isolation of individual rice chromosomes directly from a metaphase cell. Microdissection and flow sorting are two major methods for the isolation of single chromosome. These methods are dependent on the techniques of chromosome spread and chromosome suspension, respectively. In the development of this system, we avoided using chromosome spread and cell suspension was used instead. The cell wall of metaphase rice cell was cut by optical scissors. The released single chromosome was captured by an optical trap and transported to an area without cell debris. The isolated single chromosome was then collected and specific library was constructed by linker adaptor PCR. The average insert size of the library was about 300 bp. Two hundred inserts of chromosome 4 library were sequenced, and 96.5% were aligned to the corresponding sequences of rice chromosome 4. These results suggest the possible application of this method for the preparation of other subcellular structures and for the cloning of single macromolecule through a laser microbeam trap.

  15. KIF20A regulates porcine oocyte maturation and early embryo development.

    Directory of Open Access Journals (Sweden)

    Yu Zhang

    Full Text Available KIF20A (Kinesin-like family member 20A, also called mitotic kinesin-like proteins 2 (MKLP2, is a mammalian mitotic kinesin-like motor protein of the Kinesin superfamily proteins (KIFs, which was originally involved in Golgi apparatus dynamics and thought to essential for cell cycle regulation during successful cytokinesis. In the present study, we investigated whether KIF20A has roles on porcine oocyte meiotic maturation and subsequent early embryo development. By immunofluorescence staining, KIF20A was found to exhibit a dynamic localization pattern during meiosis. KIF20A was restricted to centromeres after germinal vesicle breakdown (GVBD, transferred to the midbody at telophase I (TI, and again associated with centromeres at metaphase II (MII. Inhibition of endogenous KIF20A via a specific inhibitor, Paprotrain, resulted in failure of polar body extrusion. Further cell cycle analysis showed that the percentage of oocytes that arrested at early metaphase I (MI stage increased after KIF20A activity inhibition; however, the proportion of oocytes at anaphase/telophase I (ATI and MII stages decreased significantly. Our results also showed that KIF20A inhibition did not affect spindle morphology. In addition, KIF20A was localized at the nucleus of early embryos, and KIF20A inhibition resulted in failure of early parthenogenetic embryo development. These results demonstrated that KIF20A is critical for porcine oocyte meiotic maturation and subsequent early embryo development.

  16. In vivo wound healing and antiulcer properties of white sweet potato (Ipomoea batatas).

    Science.gov (United States)

    Hermes, Daniele; Dudek, Débora N; Maria, Mariana D; Horta, Lívia P; Lima, Eliete N; de Fátima, Ângelo; Sanches, Andréia C C; Modolo, Luzia V

    2013-07-01

    The potential of tuber flour of Ipomoea batatas (L.) Lam. cv. Brazlândia Branca (white sweet potato) as wound healing and antiulcerogenic agent was investigated in vivo in animal model. Excision on the back of Wistar rats was performed to induce wounds that were topically treated with Beeler's base containing tuber flour of white sweet potato at 2.5%. Number of cells undergoing metaphase and the degree of tissue re-epithelialization were investigated 4, 7 and 10 days post-treatment. The protective effect of aqueous suspension of tuber flour (75 and 100 mg/kg animal weight) on gastric mucosa of Wistar rats was also studied by using the ethanol-induced ulceration model. Ointment based on white sweet potato at 2.5% effectively triggered the healing of cutaneous wound as attested by the increased number of cells undergoing metaphase and tissue re-epithelialization regardless the time of wound treatment. Tuber flour potentially prevented ethanol-induced gastric ulceration by suppressing edema formation and partly protecting gastric mucosa wrinkles. Crude extracts also exhibited potential as free radical scavengers. The results from animal model experiments indicate the potential of tuber flour of white sweet potato to heal wounds.

  17. Spermatogenesis and nucleolar behavior in Triatoma vandae and Triatoma williami (Hemiptera, Triatominae).

    Science.gov (United States)

    Pereira, N P; Alevi, K C C; Mendonça, P P; Azeredo-Oliveira, M T V

    2015-10-09

    This study describes spermatogenesis in Triatoma vandae and the nucleolar behavior of T. vandae and Triatoma williami, with a cytotaxonomic focus. Analysis of mitotic and meiotic metaphases of T.vandae confirms the species karyotype. T. vandae presents some characteristics during meiosis that differentiate it from T. williami, including the presence of a chromocenter with two sex chromosomes individualized during early prophase, and the presence of a bi- or tripartite corpuscle inpolyploid nuclei. It was possible to observe the compaction of chromatin during prophase resulting in holocentric chromosomes. During metaphase,the autosomes presented a ring shape and the sex chromosomes were in the center of the ring. These chromosomes were separated in anaphase. Although it is common, we did not observe the phenomenon of late migration of the sex chromosomes. By means of silver ion impregnation it was possible to describe nucleologenesis in T. vandae and T. williami. In both species we observed persistence of the nucleolar material duringmeiosis. In addition to the cells in meiotic division, we also observed the presence of polyploid nuclei in the seminiferous tubule walls that nourish the cells during cell division. The nucleolar markings reflect their capacity for synthetic activity. T. vandae and T. williami presented only one nucleolar corpuscle, which reflects low synthetic activity. This study confirms the karyotype of T. vandae, describes characteristics that differentiate T. vandae and T. williami during meiosis, and describes the phenomenon of nucleolar persistence in both species.

  18. Precocious Sister Chromatid Separation (PSCS) in Cornelia de Lange Syndrome

    Science.gov (United States)

    Kaur, Maninder; DeScipio, Cheryl; McCallum, Jennifer; Yaeger, Dinah; Devoto, Marcella; Jackson, Laird G.; Spinner, Nancy B.; Krantz, Ian D.

    2009-01-01

    The Cornelia de Lange syndrome (CdLS) (OMIM# 122470) is a dominantly inherited multisystem developmental disorder. The phenotype consists of characteristic facial features, hirsutism, abnormalities of the upper extremities ranging from subtle changes in the phalanges and metacarpal bones to oligodactyly and phocomelia, gastroesophageal dysfunction, growth retardation, and neurodevelopmental delay. Prevalence is estimated to be as high as 1 in 10,000. Recently, mutations in NIPBL were identified in sporadic and familial CdLS cases. To date, mutations in this gene have been identified in over 45% of individuals with CdLS. NIPBL is the human homolog of the Drosophila Nipped-B gene. Although its function in mammalian systems has not yet been elucidated, sequence homologs of Nipped-B in yeast (Scc2 and Mis4) are required for sister chromatid cohesion during mitosis, and a similar role was recently demonstrated for Nipped-B in Drosophila. In order to evaluate NIPBL role in sister chromatid cohesion in humans, metaphase spreads on 90 probands (40 NIPBL mutation positive and 50 NIPBL mutation negative) with CdLS were evaluated for evidence of precocious sister chromatid separation (PSCS). We screened 50 metaphases from each proband and found evidence of PSCS in 41% (compared to 9% in control samples). These studies indicate that NIPBL may play a role in sister chromatid cohesion in humans as has been reported for its homologs in Drosophila and yeast. PMID:16100726

  19. Differences in genotoxic activity of alpha-Ni3S2 on human lymphocytes from nickel-hypersensitized and nickel-unsensitized donors.

    Science.gov (United States)

    Arrouijal, F Z; Marzin, D; Hildebrand, H F; Pestel, J; Haguenoer, J M

    1992-05-01

    The genotoxic activity of alpha-Ni3S2 was assessed on human lymphocytes from nickel-hypersensitized (SSL) and nickel-unsensitized (USL) subjects. Three genotoxicity tests were performed: the sister chromatid exchange (SCE) test, the metaphase analysis test and the micronucleus test. (i) The SCE test (3-100 micrograms/ml) showed a weak but statistically significant increase in the number of SCE in both lymphocyte types with respect to controls, USL presenting a slightly higher SCE incidence but only at one concentration. (ii) The metaphase analysis test demonstrated a high dose-dependent clastogenic activity of alpha-Ni3S2 in both lymphocyte types. The frequency of chromosomal anomalies was significantly higher in USL than in SSL for all concentrations applied. (iii) The micronucleus test confirmed the dose-dependent clastogenic activity of alpha-Ni3S2 and the differences already observed between USL and SSL, i.e. the number of cells with micronuclei was statistically higher in USL. Finally, the incorporation study with alpha-63Ni3S2 showed a higher uptake of its solubilized fraction by USL. This allows an explanation of the different genotoxic action of nickel on the two cell types. In this study we demonstrated that hypersensitivity has an influence on the incorporation of alpha-Ni3S2 and subsequently on the different induction of chromosomal aberrations in human lymphocytes.

  20. Dose-response relationships for chromosome aberrations in peripheral blood lymphocytes after whole- and partial-body irradiations. Pt. 1

    International Nuclear Information System (INIS)

    Liniecki, J.; Bajerska, A.; Wyszynska, K.

    1983-01-01

    Dose-response relationships were established for yield of dicentrics and for a fraction of damaged metaphases in lymphocytes after γ-irradiation of rabbits' whole blood in vitro. These relationships were based on the scoring of cells only in their first post-stimulation division and they served as a reference system for comparison with results of 60 Co γ-irradiation in vivo, either of the whole or of predetermined parts of an animal's body. There was a statistically acceptable agreement between dose-response data established for dicentric yield after whole-body irradiation in vivo and the reference dose-response curve derived from exposure of rabbit's blood in vitro. For partial-body (1/2) irradiations there was a satisfactory agreement between the dose-response curves in vitro for dicentric yield and fraction of metaphases damaged on the one hand and the response in vivo when the latter was related to mean doses to circulating blood. However, there was a drastic disagreement with the dose responses in vitro when measured cytogenetic quantities were plotted versus mean doses to body mass. When the latter were substituted for by comparable doses to circulating blood the in vivo-in vitro agreement was acceptable after irradiation. (orig.)

  1. Development of software and modification of Q-FISH protocol for estimation of individual telomere length in immunopathology.

    Science.gov (United States)

    Barkovskaya, M Sh; Bogomolov, A G; Knauer, N Yu; Rubtsov, N B; Kozlov, V A

    2017-04-01

    Telomere length is an important indicator of proliferative cell history and potential. Decreasing telomere length in the cells of an immune system can indicate immune aging in immune-mediated and chronic inflammatory diseases. Quantitative fluorescent in situ hybridization (Q-FISH) of a labeled (C 3 TA[Formula: see text] peptide nucleic acid probe onto fixed metaphase cells followed by digital image microscopy allows the evaluation of telomere length in the arms of individual chromosomes. Computer-assisted analysis of microscopic images can provide quantitative information on the number of telomeric repeats in individual telomeres. We developed new software to estimate telomere length. The MeTeLen software contains new options that can be used to solve some Q-FISH and microscopy problems, including correction of irregular light effects and elimination of background fluorescence. The identification and description of chromosomes and chromosome regions are essential to the Q-FISH technique. To improve the quality of cytogenetic analysis after Q-FISH, we optimized the temperature and time of DNA-denaturation to get better DAPI-banding of metaphase chromosomes. MeTeLen was tested by comparing telomere length estimations for sister chromatids, background fluorescence estimations, and correction of nonuniform light effects. The application of the developed software for analysis of telomere length in patients with rheumatoid arthritis was demonstrated.

  2. Department of Radiation and Environmental Biology - Overview

    International Nuclear Information System (INIS)

    Cebulska-Wasilewska, A.

    2000-01-01

    Full text:The year 1999 we devoted mainly to the activities concerning our basic research, and requirements and expectations of three research projects. The environmental project from the European Community was supporting our research in the issues of human monitoring of occupational exposure to pesticides. The two other radiobiology projects from the State Committee of Research were supporting our search on the biological efficiency and its enhancement of radio-therapeutic sources of various LET radiation. We succeeded fruitful co-operation with colleagues from Academy of Mining and Metallurgy that let us go faster with modernization of our laboratory by automation of our methods for screening cytogenetic damages. A lot of efforts were paid to modify our work by automatic reports of the coordinates of aberrant metaphases, and to make a smooth work of our new and own metaphase finder. We are sure that our new and unique research tool will not only enhance the accuracy and speed of measurements, but will also be useful for the purpose of the retrospective biological dosimetry of absorbed doses. We have applied fluorescent in situ hybridization (FISH) for cytogenetic studies of biological effects induced by neutrons. Now, we are looking forward to apply this technique in a combination with the DNA damage measures done by SCGE assay, to our research on mechanisms of the induction and repair, or interaction of the lesions induced by genotoxic agents. Understanding of the regulation of these processes could be a good goal for the new century to come. (author)

  3. Structural Chromosomal Alterations Induced by Dietary Bioflavonoids in Fanconi Anemia Lymphocytes

    Directory of Open Access Journals (Sweden)

    Gonzalo Guevara

    2007-06-01

    Full Text Available IntroductionFanconi anemia is an autosomal recessive diseasecharacterized by a variety of congenital abnormalities,progressive bone marrow failure,increased chromosomal instability and higherrisk to acute myeloid leukemia, solid tumors. Thisentity can be considered an appropriate biologicalmodel to analyze natural substances with possiblegenotoxic effect. The aims of this study wereto describe and quantify structural chromosomalaberrations induced by 5 flavones, 2 isoflavonesand a topoisomerase II chemotherapeutic inhibitorin Fanconi anemia lymphocytes in order todetermine chromosomal numbers changes and/or type of chromosomal damage.Materials and methodsChromosomes stimulated by phytohaemagglutininM, from Fanconi anemia lymphocytes,were analysed by conventional cytogenetic culture.For each chemical substance and controls,one hundred metaphases were evaluated. Chromosomalalterations were documented by photographyand imaging analyzer. To statisticalanalysis was used chi square test to identify significantdifferences between frequencies of chromosomaldamage of basal and exposed cellcultured a P value less than 0.05.ResultsThere were 431 chromosomal alterations in1000 metaphases analysed; genistein was themore genotoxic bioflavonoid, followed in descendentorder by genistin, fisetin, kaempferol,quercetin, baicalein and miricetin. Chromosomalaberrations observed were: chromatidbreaks, chromosomal breaks, cromatid andchromosomal gaps, quadriratials exchanges,dicentrics chromosome and complex rearrangements.ConclusionBioflavonoids as genistein, genistin and fisetin,which are commonly present in the human diet,showed statistical significance in the number ofchromosomal aberrations in Fanconi anemialymphocytes, regarding the basal damage.

  4. Loss of maternal ATRX results in centromere instability and aneuploidy in the mammalian oocyte and pre-implantation embryo.

    Directory of Open Access Journals (Sweden)

    Claudia Baumann

    2010-09-01

    Full Text Available The α-thalassemia/mental retardation X-linked protein (ATRX is a chromatin-remodeling factor known to regulate DNA methylation at repetitive sequences of the human genome. We have previously demonstrated that ATRX binds to pericentric heterochromatin domains in mouse oocytes at the metaphase II stage where it is involved in mediating chromosome alignment at the meiotic spindle. However, the role of ATRX in the functional differentiation of chromatin structure during meiosis is not known. To test ATRX function in the germ line, we developed an oocyte-specific transgenic RNAi knockdown mouse model. Our results demonstrate that ATRX is required for heterochromatin formation and maintenance of chromosome stability during meiosis. During prophase I arrest, ATRX is necessary to recruit the transcriptional regulator DAXX (death domain associated protein to pericentric heterochromatin. At the metaphase II stage, transgenic ATRX-RNAi oocytes exhibit abnormal chromosome morphology associated with reduced phosphorylation of histone 3 at serine 10 as well as chromosome segregation defects leading to aneuploidy and severely reduced fertility. Notably, a large proportion of ATRX-depleted oocytes and 1-cell stage embryos exhibit chromosome fragments and centromeric DNA-containing micronuclei. Our results provide novel evidence indicating that ATRX is required for centromere stability and the epigenetic control of heterochromatin function during meiosis and the transition to the first mitosis.

  5. Cytomictic Anomalous Male Meiosis and 2n Pollen Grain Formation in Mertensia echioides Benth. (Boraginaceae from Kashmir Himalaya

    Directory of Open Access Journals (Sweden)

    Reyaz Ahmad Malik

    2014-01-01

    Full Text Available Presently Mertensia echioides Benth. (Boraginaceae collected from Kashmir Himalaya, India, is cytologically analyzed for the first time revealing 2n=2x=24 (diploid. Interestingly we found 4.3–6.2% syncytic meiocytes/PMCs with 2n=4x=48 (tetraploid in addition to normal meiocytes (2n=24 during male meiosis. These comparatively larger PMCs (pollen mother cells lead to the formation of fertile giant 2n pollen grains. A frequency of 6.4–13.3% PMCs shows transfer of chromatin material at prophase-I and, therefore, results in aneuploid meiocytes. Whole chromatin transfer by the process of cytomixis could also have led to the formation of tetraploid cells. Translocation heterozygosity is also evident in the form of multivalents in 12–17% diploid (2x meiocytes at diakinesis and metaphase-I and is reported for the first time in this species. The syncytes formed depict open chain hexavalent and quadrivalent formation in the three populations with different frequencies. Moreover chromatin stickiness at metaphase-I is observed in 45% of PMCs in population-1 (P-1. Syncyte or unreduced PMC formation leading to unreduced fertile gametes is here speculated to act as a possible way out for infraspecific polyploidization in the species.

  6. Phosphorylation of the Ndc80 complex protein, HEC1, by Nek2 kinase modulates chromosome alignment and signaling of the spindle assembly checkpoint.

    Science.gov (United States)

    Wei, Randy; Ngo, Bryan; Wu, Guikai; Lee, Wen-Hwa

    2011-10-01

    The spindle assemble checkpoint (SAC) is critical for accurate chromosome segregation. Hec1 contributes to chromosome segregation in part by mediating SAC signaling and chromosome alignment. However, the molecular mechanism by which Hec1 modulates checkpoint signaling and alignment remains poorly understood. We found that Hec1 serine 165 (S165) is preferentially phosphorylated at kinetochores. Phosphorylated Hec1 serine 165 (pS165) specifically localized to kinetochores of misaligned chromosomes, showing a spatiotemporal distribution characteristic of SAC molecules. Expressing an RNA interference (RNAi)-resistant S165A mutant in Hec1-depleted cells permitted normal progression to metaphase, but accelerated the metaphase-to-anaphase transition. The S165A cells were defective in Mad1 and Mad2 localization to kinetochores, regardless of attachment status. These cells often entered anaphase with lagging chromosomes and elicited increased segregation errors and cell death. In contrast, expressing S165E mutant in Hec1-depleted cells triggered defective chromosome alignment and severe mitotic arrest associated with increased Mad1/Mad2 signals at prometaphase kinetochores. A small portion of S165E cells eventually bypassed the SAC but showed severe segregation errors. Nek2 is the primary kinase responsible for kinetochore pS165, while PP1 phosphatase may dephosphorylate pS165 during SAC silencing. Taken together, these results suggest that modifications of Hec1 S165 serve as an important mechanism in modulating SAC signaling and chromosome alignment.

  7. Sensitivity of the meiotic stage to hyperthermia during in vitro maturation of porcine oocytes.

    Science.gov (United States)

    Nishio, Katsutoshi; Yamazaki, Mado; Taniguchi, Masayasu; Besshi, Kazuhiko; Morita, Fumio; Kunihara, Toshiki; Tanihara, Fuminori; Takemoto, Tatsuya; Otoi, Takeshige

    2017-03-01

    The present study was conducted to clarify whether the meiotic stage of porcine oocytes has the highest sensitivity to hyperthermia during in vitro maturation by evaluating meiotic competence and DNA damage. Oocytes were exposed to 41 °C for 12 h at various intervals during 48 h of maturation culture. When the oocytes were exposed to 41 °C from 12 to 24 h of the maturation culture, the proportion of oocytes reaching metaphase II (MII) decreased as compared to the control oocytes cultured at 38.5 °C (P < 0.05). Moreover, the proportions of DNA fragmentation in all oocytes exposed to 41 °C in each culture period after 12 h from the start of maturation culture were significantly higher (P < 0.05) than for the control oocytes. When the meiotic stage of oocytes cultured at 38.5 °C between 12 and 24 h was examined, the majority of oocytes remained at the germinal vesicle (GV) stage at 12 h and approximately half of the oocytes reached metaphase I (MI) at 24 h. These results indicate that the meiotic stage of porcine oocytes having the highest sensitivity to hyperthermia during in vitro maturation is a transition period from the GV stage to the MI stage.

  8. Cloning and comparative mapping of a human chromosome 4-specific alpha satellite DNA sequence

    Energy Technology Data Exchange (ETDEWEB)

    D' Aiuto, L.; Marzella, R.; Archidiacono, N.; Rocchi, M. (Universita di Bari (Italy)); Antonacci, R. (Instituto Anatomia Umana Normale, Modena (Italy))

    1993-11-01

    The authors have isolated and characterized two human alphoid DNA clones: p4n1/4 and pZ4.1. Clone p4n1/4 identifies specifically the centromeric region of chromosome 4; pZ4.1 recognizes a subset of alphoid DNA shared by chromosomes 4 and 9. The specificity was determined using fluorescence in situ hybridization experiments on metaphase spreads and Southern blotting analysis of human-hamster somatic cell hybrids. The genomic organization of both subsets was also investigated. Comparative mapping on chimpanzee and gorilla chromosomes was performed. p4n1/4 hybridizes to chimpanzee chromosomes 11 and 13, homologs of human chromosomes 9 and 2q, respectively. On gorilla metaphase spreads, p4n1/4 hybridizes exclusively to the centromeric region of chromosome 19, partially homologous to human chromosome 17. No hybridization signal was detected on chromosome 3 of both chimpanzee and gorilla, in both species homolog of human chromosome 4. Identical comparative mapping results were obtained using pZ4.1 probe, although the latter recognizes an alphoid subset distinct from the one recognized by p4n1/4. The implications of these results in the evolution of centromeric regions of primate chromosomes are discussed. 33 refs., 4 figs.

  9. A de novo chromosomal abnormality in Cri du Chat syndrome.

    Science.gov (United States)

    Sun, Shunchang C; Luo, Fuwei W; Zhou, Zhiming M; Peng, Yunsheng S; Song, Huiwen W

    2014-07-01

    To find the length and location of the deletions in the short arm of chromosome 5 in one case of Cri du Chat syndrome using oligo array comparative genomic hybridization. Metaphase chromosomes were prepared from peripheral blood lymphocyte cultures using standard cytogenetic protocols. Chromosomal analysis was done in G-banded metaphases. Oligo array comparative genomic hybridization and fluorescence in situ hybridization were performed by the commercially available kits. Oligonucleotide array comparative genomic hybridization (CGH) analysis revealed a 23.263 Mb deletion at region 5p14.2-->qter, combined with a duplication of 14.602 Mb in size in the area 12p13.1-->pter. Chromosomal aberrations were confirmed by fluorescence in situ hybridization. The male neonate with Cri du Chat syndrome had an unbalanced translocation which was inherited from his father who was a balanced carrier with a karyotype 46, XY, t (5; 12) (p14.2; p13.1). This report shows the clinical utility of the oligonucleotide array in the detection of submicroscopic chromosomal aberrations, thus improving the molecular diagnosis of Cri du Chat syndrome.

  10. Cytogenetic data on the agro-predatory ant Megalomyrmex incisus Smith, 1947 and its host, Mycetophylax conformis (Mayr, 1884 (Hymenoptera, Formicidae

    Directory of Open Access Journals (Sweden)

    Danon Clemes Cardoso

    2017-01-01

    Full Text Available We provide the first karyotype description of the agro-predatory ant species Megalomyrmex incisus Smith, 1947 (Myrmicinae, Formicidae, and chromosome counts of its host Mycetophylax conformis (Mayr, 1884 (Myrmicinae, Formicidae from geographically distinct populations. Colonies of both species were sampled from coastal areas of Ilhéus, Bahia, Brazil, and transferred to the laboratory. Metaphase spreads were prepared from the cerebral ganglia of defecated larvae. The slides were examined and pictures of the best metaphases were taken. The chromosome number for M. incisus was 2n=50 and n=25. The karyotype of this species consists of 20 metacentric and 5 submetacentric pairs. Thus, the karyotype formula of the diploid set was 2K=40M + 10SM and a fundamental number FN=100. The host species M. conformis has 2n=30 and the karyotype consisting of 11 metacentric and 4 submetacentric pairs. The karyotype formula was 2K=22M + 8SM, and a fundamental number FN=60. M. incisus showed a slightly higher chromosome number, placed at the marginal range of the known distribution of haploid karyotypes of the Myrmicinae. The chromosome number and chromosomal morphology of M. conformis corresponded to those of previously studied populations, suggesting its karyotype stability.

  11. Unconventional actin conformations localize on intermediate filaments in mitosis

    International Nuclear Information System (INIS)

    Hubert, Thomas; Vandekerckhove, Joel; Gettemans, Jan

    2011-01-01

    Research highlights: → Unconventional actin conformations colocalize with vimentin on a cage-like structure in metaphase HEK 293T cells. → These conformations are detected with the anti-actin antibodies 1C7 ('lower dimer') and 2G2 ('nuclear actin'), but not C4 (monomeric actin). → Mitotic unconventional actin cables are independent of filamentous actin or microtubules. → Unconventional actin colocalizes with vimentin on a nocodazole-induced perinuclear dense mass of cables. -- Abstract: Different structural conformations of actin have been identified in cells and shown to reside in distinct subcellular locations of cells. In this report, we describe the localization of actin on a cage-like structure in metaphase HEK 293T cells. Actin was detected with the anti-actin antibodies 1C7 and 2G2, but not with the anti-actin antibody C4. Actin contained in this structure is independent of microtubules and actin filaments, and colocalizes with vimentin. Taking advantage of intermediate filament collapse into a perinuclear dense mass of cables when microtubules are depolymerized, we were able to relocalize actin to such structures. We hypothesize that phosphorylation of intermediate filaments at mitosis entry triggers the recruitment of different actin conformations to mitotic intermediate filaments. Storage and partition of the nuclear actin and antiparallel 'lower dimer' actin conformations between daughter cells possibly contribute to gene transcription and transient actin filament dynamics at G1 entry.

  12. Interphase Chromosome Profiling: A Method for Conventional Banded Chromosome Analysis Using Interphase Nuclei.

    Science.gov (United States)

    Babu, Ramesh; Van Dyke, Daniel L; Dev, Vaithilingam G; Koduru, Prasad; Rao, Nagesh; Mitter, Navnit S; Liu, Mingya; Fuentes, Ernesto; Fuentes, Sarah; Papa, Stephen

    2018-02-01

    - Chromosome analysis on bone marrow or peripheral blood samples fails in a small proportion of attempts. A method that is more reliable, with similar or better resolution, would be a welcome addition to the armamentarium of the cytogenetics laboratory. - To develop a method similar to banded metaphase chromosome analysis that relies only on interphase nuclei. - To label multiple targets in an equidistant fashion along the entire length of each chromosome, including landmark subtelomere and centromere regions. Each label so generated by using cloned bacterial artificial chromosome probes is molecularly distinct with unique spectral characteristics, so the number and position of the labels can be tracked to identify chromosome abnormalities. - Interphase chromosome profiling (ICP) demonstrated results similar to conventional chromosome analysis and fluorescence in situ hybridization in 55 previously studied cases and obtained useful ICP chromosome analysis results on another 29 cases in which conventional methods failed. - ICP is a new and powerful method to karyotype peripheral blood and bone marrow aspirate preparations without reliance on metaphase chromosome preparations. It will be of particular value for cases with a failed conventional analysis or when a fast turnaround time is required.

  13. A Role for the Chaperone Complex BAG3-HSPB8 in Actin Dynamics, Spindle Orientation and Proper Chromosome Segregation during Mitosis

    Science.gov (United States)

    Fuchs, Margit; Lambert, Herman; Jetté, Alexandra; Elowe, Sabine; Landry, Jacques; Lavoie, Josée N.

    2015-01-01

    The co-chaperone BAG3, in complex with the heat shock protein HSPB8, plays a role in protein quality control during mechanical strain. It is part of a multichaperone complex that senses damaged cytoskeletal proteins and orchestrates their seclusion and/or degradation by selective autophagy. Here we describe a novel role for the BAG3-HSPB8 complex in mitosis, a process involving profound changes in cell tension homeostasis. BAG3 is hyperphosphorylated at mitotic entry and localizes to centrosomal regions. BAG3 regulates, in an HSPB8-dependent manner, the timely congression of chromosomes to the metaphase plate by influencing the three-dimensional positioning of the mitotic spindle. Depletion of BAG3 caused defects in cell rounding at metaphase and dramatic blebbing of the cortex associated with abnormal spindle rotations. Similar defects were observed upon silencing of the autophagic receptor p62/SQSTM1 that contributes to BAG3-mediated selective autophagy pathway. Mitotic cells depleted of BAG3, HSPB8 or p62/SQSTM1 exhibited disorganized actin-rich retraction fibres, which are proposed to guide spindle orientation. Proper spindle positioning was rescued in BAG3-depleted cells upon addition of the lectin concanavalin A, which restores cortex rigidity. Together, our findings suggest the existence of a so-far unrecognized quality control mechanism involving BAG3, HSPB8 and p62/SQSTM1 for accurate remodelling of actin-based mitotic structures that guide spindle orientation. PMID:26496431

  14. Genomic instability in rat: Breakpoints induced by ionising radiation and interstitial telomeric-like sequences

    Energy Technology Data Exchange (ETDEWEB)

    Camats, Nuria [Institut de Biotecnologia i Biomedicina (IBB), Universitat Autonoma de Barcelona, 08193 Barcelona (Spain); Departament de Biologia Cel.lular, Fisiologia i Immunologia Universitat Autonoma de Barcelona, 08193 Barcelona (Spain); Ruiz-Herrera, Aurora [Departament de Biologia Cel.lular, Fisiologia i Immunologia Universitat Autonoma de Barcelona, 08193 Barcelona (Spain); Parrilla, Juan Jose [Servicio de Ginecologia y Obstetricia, Hospital Universitario Virgen de la Arrixaca, Ctra, Madrid-Cartagena, s/n, El Palmar, 30120 Murcia (Spain); Acien, Maribel [Servicio de Ginecologia y Obstetricia, Hospital Universitario Virgen de la Arrixaca, Ctra, Madrid-Cartagena, s/n, El Palmar, 30120 Murcia (Spain); Paya, Pilar [Servicio de Ginecologia y Obstetricia, Hospital Universitario Virgen de la Arrixaca, Ctra, Madrid-Cartagena, s/n, El Palmar, 30120 Murcia (Spain); Giulotto, Elena [Dipartimento di Genetica e Microbiologia Adriano Buzzati Traverso, Universita degli Studi di Pavia, 27100 Pavia (Italy); Egozcue, Josep [Departament de Biologia Cel.lular, Fisiologia i Immunologia Universitat Autonoma de Barcelona, 08193 Barcelona (Spain); Garcia, Francisca [Institut de Biotecnologia i Biomedicina (IBB), Universitat Autonoma de Barcelona, 08193 Barcelona (Spain); Garcia, Montserrat [Institut de Biotecnologia i Biomedicina (IBB), Universitat Autonoma de Barcelona, 08193 Barcelona (Spain) and Departament de Biologia Cellular, Fisiologia i Immunologia Universitat Autonoma de Barcelona, 08193 Barcelona (Spain)]. E-mail: Montserrat.Garcia.Caldes@uab.es

    2006-03-20

    The Norwegian rat (Rattus norvegicus) is the most widely studied experimental species in biomedical research although little is known about its chromosomal structure. The characterisation of possible unstable regions of the karyotype of this species would contribute to the better understanding of its genomic architecture. The cytogenetic effects of ionising radiation have been widely used for the study of genomic instability, and the importance of interstitial telomeric-like sequences (ITSs) in instability of the genome has also been reported in previous studies in vertebrates. In order to describe the unstable chromosomal regions of R. norvegicus, the distribution of breakpoints induced by X-irradiation and ITSs in its karyotype were analysed in this work. For the X-irradiation analysis, 52 foetuses (from 14 irradiated rats) were studied, 4803 metaphases were analysed, and a total of 456 breakpoints induced by X-rays were detected, located in 114 chromosomal bands, with 25 of them significantly affected by X-irradiation (hot spots). For the analysis of ITSs, three foetuses (from three rats) were studied, 305 metaphases were analysed and 121 ITSs were detected, widely distributed in the karyotype of this species. Seventy-six percent of all hot spots analysed in this study were co-localised with ITSs.

  15. The Relationship Between Transcript Expression Levels of Nuclear Encoded (TFAM, NRF1 and Mitochondrial Encoded (MT-CO1 Genes in Single Human Oocytes During Oocyte Maturation

    Directory of Open Access Journals (Sweden)

    Ghaffari Novin M.

    2015-06-01

    Full Text Available In some cases of infertility in women, human oocytes fail to mature when they reach the metaphase II (MII stage. Mitochondria plays an important role in oocyte maturation. A large number of mitochondrial DNA (mtDNA, copied in oocytes, is essential for providing adenosine triphosphate (ATP during oocyte maturation. The purpose of this study was to identify the relationship between transcript expression levels of the mitochondrial encoded gene (MT-CO1 and two nuclear encoded genes, nuclear respiratory factor 1 (NRF1 and mitochondrial transcription factor A (TFAM in various stages of human oocyte maturation. Nine consenting patients, age 21-35 years old, with male factors were selected for ovarian stimulation and intracytoplasmic sperm injection (ICSI procedures. mRNA levels of mitochondrial- related genes were performed by singlecell TaqMan® quantitative real-time polymerase chain reaction (qRT-PCR. There was no significant relationship between the relative expression levels in germinal vesicle (GV stage oocytes (p = 0.62. On the contrary, a significant relationship was seen between the relative expression levels of TFAM and NRF1 and the MT-CO1 genes at the stages of metaphase I (MI and MII (p = 0.03 and p = 0.002. A relationship exists between the transcript expression levels of TFAM and NRF1, and MT-CO1 genes in various stages of human oocyte maturation.

  16. Biosynthesis of copper nanoparticles and its effect on actively dividing cells of mitosis in Allium cepa.

    Science.gov (United States)

    Nagaonkar, Dipali; Shende, Sudhir; Rai, Mahendra

    2015-01-01

    Nanobiotechnological application of copper nanoparticles has paved the way for advancement in agriculture owing to its bactericidal and fungicidal activities. Recently, researchers have focussed on bioinspired synthesis of copper nanoparticles as a viable alternative to existing physicochemical techniques. For the commercialization of nanocopper, the toxicity evaluation is a major issue. In this context, Citrus medica (L.) fruit extract-mediated copper nanoparticles were synthesized and its different concentrations (10, 20, 40, 60, 80, and 100 µg mL(-1) ) were evaluated for its effect on actively dividing cells of Allium cepa. The study clearly revealed that copper nanoparticles increased mitotic index up to the concentration of 20 µg mL(-1) . In addition, a gradual decline in mitotic index and increase in abnormality index was observed as the concentration of copper nanoparticles and treatment duration were increased. Aberrations in chromosomal behavior such as sticky and disturbed chromosomes in metaphase and anaphase, c-metaphase, bridges, laggard, disturbed telophase, and vacuolated nucleus were also observed. © 2015 American Institute of Chemical Engineers.

  17. Genomic alterations detected by comparative genomic hybridization in ovarian endometriomas

    Directory of Open Access Journals (Sweden)

    L.C. Veiga-Castelli

    2010-08-01

    Full Text Available Endometriosis is a complex and multifactorial disease. Chromosomal imbalance screening in endometriotic tissue can be used to detect hot-spot regions in the search for a possible genetic marker for endometriosis. The objective of the present study was to detect chromosomal imbalances by comparative genomic hybridization (CGH in ectopic tissue samples from ovarian endometriomas and eutopic tissue from the same patients. We evaluated 10 ovarian endometriotic tissues and 10 eutopic endometrial tissues by metaphase CGH. CGH was prepared with normal and test DNA enzymatically digested, ligated to adaptors and amplified by PCR. A second PCR was performed for DNA labeling. Equal amounts of both normal and test-labeled DNA were hybridized in human normal metaphases. The Isis FISH Imaging System V 5.0 software was used for chromosome analysis. In both eutopic and ectopic groups, 4/10 samples presented chromosomal alterations, mainly chromosomal gains. CGH identified 11q12.3-q13.1, 17p11.1-p12, 17q25.3-qter, and 19p as critical regions. Genomic imbalances in 11q, 17p, 17q, and 19p were detected in normal eutopic and/or ectopic endometrium from women with ovarian endometriosis. These regions contain genes such as POLR2G, MXRA7 and UBA52 involved in biological processes that may lead to the establishment and maintenance of endometriotic implants. This genomic imbalance may affect genes in which dysregulation impacts both eutopic and ectopic endometrium.

  18. Karyotypes, constitutive heterochromatin, and genomic DNA values in the blowfly genera Chrysomya, Lucilia, and Protophormia (Diptera: Calliphoridae).

    Science.gov (United States)

    Ullerich, Fritz-Helmut; Schöttke, Michael

    2006-06-01

    The karyotypes and C-banding patterns of Chrysomya species C. marginalis, C. phaonis, C. pinguis, C. saffranea, C. megacephala (New Guinean strain), Lucilia sericata, and Protophormia terraenovae are described. All species are amphogenic and have similar chromosome complements (2n = 12), including an XY-XX sex-chromosome pair varying in size and morphology between species. Additionally, the C-banding pattern of the monogenic species Chrysomya albiceps is presented. The DNA contents of these and of further species Chrysomya rufifacies, Chrysomya varipes, and Chrysomya putoria were assessed on mitotic metaphases by Feulgen cytophotometry. The average 2C DNA value of the male genomes ranged from 1.04 pg in C. varipes to 2.31 pg in C. pinguis. The DNA content of metaphase X chromosomes varied from 0.013 pg (= 1.23% of the total genome) in C. varipes to 0.277 pg (12.20%) in L. sericata; that of Y chromosomes ranged from 0.003 pg (0.27%) in C. varipes to 0.104 pg (5.59%) in L. sericata. In most species, the corresponding 5 large chromosome pairs showed similar relative DNA contents. The data suggest that the interspecific DNA differences in most species are mainly due to quantitative variation of (repetitive) sequences lying outside the centromeric heterochromatin blocks of the large chromosomes. The results are also discussed with regard to phylogenetic relationships of some species.

  19. X-ray-induced dicentric yields in lymphocytes of the teleost, Umbra limi

    Energy Technology Data Exchange (ETDEWEB)

    Suyama, I. (National Inst. of Radiological Sciences, Chiba (Japan). Div. of Environmental Health); Etoh, H. (National Inst. of Radiological Sciences, Chiba (Japan). Div. of Biology)

    1983-01-01

    A microculture technique was applied to the study of lymphocytes of Umbra limi, which have a low number of large meta- and submetacentric chromosomes (2n=22). On the 5th day (90 h) and later after initiation of culture at 20/sup 0/C, some cultures provided well spread metaphase chromosomes for analyses. After initiation, cultures were irradiated with 50, 100, 150 and 200 R of 200 kVp X-rays. The cultures were harvested on the 5th day, at which time all arrested metaphase chromosomes were in their first division. The dicentric yields induced in X-irradiated Umbra lymphocytes were observed to be significantly (P=0.05) lower than those in human lymphocytes. The resulting dose-response relationship for dicentric yield was described by the quadratic equation Y=aD+bD/sup 2/. The advantage of this method lies in the fact that small amounts (0.1 ml) of blood can be repeatedly withdrawn from the fish after a minimal interval of 2 weeks.

  20. X-ray-induced dicentric yields in lymphocytes of the teleost, Umbra limi

    International Nuclear Information System (INIS)

    Suyama, I.; Etoh, H.

    1983-01-01

    A microculture technique was applied to the study of lymphocytes of Umbra limi, which have a low number of large meta- and submetacentric chromosomes (2n=22). On the 5th day (90 h) and later after initiation of culture at 20 0 C, some cultures provided well spread metaphase chromosomes for analyses. After initiation, cultures were irradiated with 50, 100, 150 and 200 R of 200 kVp X-rays. The cultures were harvested on the 5th day, at which time all arrested metaphase chromosomes were in their first division. The dicentric yields induced in X-irradiated Umbra lymphocytes were observed to be significantly (P=0.05) lower than those in human lymphocytes. The resulting dose-response relationship for dicentric yield was described by the quadratic equation Y=aD+bD 2 . The advantage of this method lies in the fact that small amounts (0.1 ml) of blood can be repeatedly withdrawn from the fish after a minimal interval of 2 weeks. (orig./AJ)

  1. Morphological images analysis and chromosomic aberrations classification based on fuzzy logic

    International Nuclear Information System (INIS)

    Souza, Leonardo Peres

    2011-01-01

    This work has implemented a methodology for automation of images analysis of chromosomes of human cells irradiated at IEA-R1 nuclear reactor (located at IPEN, Sao Paulo, Brazil), and therefore subject to morphological aberrations. This methodology intends to be a tool for helping cytogeneticists on identification, characterization and classification of chromosomal metaphasic analysis. The methodology development has included the creation of a software application based on artificial intelligence techniques using Fuzzy Logic combined with image processing techniques. The developed application was named CHRIMAN and is composed of modules that contain the methodological steps which are important requirements in order to achieve an automated analysis. The first step is the standardization of the bi-dimensional digital image acquisition procedure through coupling a simple digital camera to the ocular of the conventional metaphasic analysis microscope. Second step is related to the image treatment achieved through digital filters application; storing and organization of information obtained both from image content itself, and from selected extracted features, for further use on pattern recognition algorithms. The third step consists on characterizing, counting and classification of stored digital images and extracted features information. The accuracy in the recognition of chromosome images is 93.9%. This classification is based on classical standards obtained at Buckton [1973], and enables support to geneticist on chromosomic analysis procedure, decreasing analysis time, and creating conditions to include this method on a broader evaluation system on human cell damage due to ionizing radiation exposure. (author)

  2. Par1b links lumen polarity with LGN-NuMA positioning for distinct epithelial cell division phenotypes.

    Science.gov (United States)

    Lázaro-Diéguez, Francisco; Cohen, David; Fernandez, Dawn; Hodgson, Louis; van Ijzendoorn, Sven C D; Müsch, Anne

    2013-10-28

    Columnar epithelia establish their luminal domains and their mitotic spindles parallel to the basal surface and undergo symmetric cell divisions in which the cleavage furrow bisects the apical domain. Hepatocyte lumina interrupt the lateral domain of neighboring cells perpendicular to two basal domains and their cleavage furrow rarely bifurcates the luminal domains. We determine that the serine/threonine kinase Par1b defines lumen position in concert with the position of the astral microtubule anchoring complex LGN-NuMA to yield the distinct epithelial division phenotypes. Par1b signaling via the extracellular matrix (ECM) in polarizing cells determined RhoA/Rho-kinase activity at cell-cell contact sites. Columnar MDCK and Par1b-depleted hepatocytic HepG2 cells featured high RhoA activity that correlated with robust LGN-NuMA recruitment to the metaphase cortex, spindle alignment with the substratum, and columnar organization. Reduced RhoA activity at the metaphase cortex in HepG2 cells and Par1b-overexpressing MDCK cells correlated with a single or no LGN-NuMA crescent, tilted spindles, and the development of lateral lumen polarity.

  3. Comparative Genomic Hybridization of Human Malignant Gliomas Reveals Multiple Amplification Sites and Nonrandom Chromosomal Gains and Losses

    Science.gov (United States)

    Schròck, Evelin; Thiel, Gundula; Lozanova, Tanka; du Manoir, Stanislas; Meffert, Marie-Christine; Jauch, Anna; Speicher, Michael R.; Nürnberg, Peter; Vogel, Siegfried; Janisch, Werner; Donis-Keller, Helen; Ried, Thomas; Witkowski, Regine; Cremer, Thomas

    1994-01-01

    Nine human malignant gliomas (2 astrocytomas grade III and 7 glioblastomas) were analyzed using comparative genomic hybridization (CGH). In addition to the amplification of the EGFR gene at 7p12 in 4 of 9 cases, six new amplification sites were mapped to 1q32, 4q12, 7q21.1, 7q21.2-3, 12p, and 22q12. Nonrandom chromosomal gains and losses were identified with overrepresentation of chromosome 7 and underrepresentation of chromosome 10 as the most frequent events (1 of 2 astrocytomas, 7 of 7 glioblastomas). Gain of a part or the whole chromosome 19 and losses of chromosome bands 9pter-23 and 22q13 were detected each in five cases. Loss of chromosome band 17p13 and gain of chromosome 20 were revealed each in three cases. The validity of the CGH data was confirmed using interphase cytogenetics with YAC clones, chromosome painting in tumor metaphase spreads, and DNA fingerprinting. A comparison of CGH data with the results of chromosome banding analyses indicates that metaphase spreads accessible in primary tumor cell cultures may not represent the clones predominant in the tumor tissue ImagesFigure 1Figure 4Figure 6 PMID:8203461

  4. Results of cytogenetic surveillance of rural populations adjoining to Semipalatinsk test site

    International Nuclear Information System (INIS)

    Abdil'dinova, G.Zh.; Kundakbaeva, G.B.; Zhunusova, A.B.

    1997-01-01

    Purpose of the work is study of somatic cytogenetic effects with taking into consideration chromosomal aberration of population from Beskaragaj district of Semipalatinsk region exposed to influence of conducted nuclear tests on Semipalatinsk test site. Cultures of lymphocyates of peripheral flood were studied. Control group made up 15 man living in Akmola region locating out of radiation contaminated zone. It is determined, that common frequency of chromosomal aberration make up 3.6 on 100 cells. At that frequency of pair fragments made up 2.0 on 100 metaphases (0.2 in control group); dicentrics and center rings were revealed with summary frequency 0.6 on 100 metaphases and that is approximately in 15 fold greater than control index - 0.03 on 100 cells. Stable radiation markers (translocations, deletions) revealed with frequency 1.07 on 100 cells (control make up 0.4 on 100 cells). By results of of conducted cytogenetic analysis the attempt of mean group effective equivalent dose reconstruction received with inhabitants of Beskaragaj district is undertaken. It was calculated, that population of researched district have been got dose loading in 9.09 rem

  5. Cytogenetic study in workers occupationally exposed to mercury fulminate.

    Science.gov (United States)

    Anwar, W A; Gabal, M S

    1991-05-01

    This study was conducted to evaluate the cytogenetic effects in male workers exposed to mercury fulminate. A total of 29 male workers and 29 age- and sex-matched controls were examined. The mean mercury level in urine from the exposed workers was 123.2 +/- 54.1 micrograms/l compared with 39.2 +/- 11.1 micrograms/l in the control group. The difference was statistically significant (P less than 0.001). Metaphase chromosomes were studied. Micronucleated peripheral blood lymphocytes were also analyzed in cytochalasin B blocked binucleated lymphocytes. The percentage of metaphases with chromosomal aberrations was significantly higher (P less than 0.001) in the exposed group (6.1 +/- 2.3) compared to the control group (2.8 +/- 0.7). The chromosomal aberrations detected were in the form of gaps, breaks and fragments. A significant increase in the incidence of micronucleated lymphocytes was found among the exposed group (7.1 +/- 4.2) compared to the control group (5.4 +/- 2.2) (P less than 0.01). The detected chromosomal damage correlated neither with the duration of exposure nor with the urinary mercury level.

  6. Analysis of chromosome lesions in women with gynecological disorders, residing in territories contaminated as a result of the Chernobyl accident

    International Nuclear Information System (INIS)

    Ivanova, T. I.; Kondrashova, T. V.; Krikunova, L. I.; Shenterva, N. I.; Song, Jie Young; Han, Young Soo; Yun, Yeon Sook

    2004-01-01

    Spontaneous levels of chromosome aberrations in peripheral lymphocytes of 95 women were analyzed in the course of medical examination of residents of Bryansk region territories (Klintsy district, and the town of Klintsy) contaminated after the Chernobyl NPP accident. All recognizable chromosome lesion types were scored in the first in vitro division metaphases stained with azure-eosin. The mean total aberration frequency in the sample studied was 5.1±0.4 per 100 metaphases, the main contribution being made by chromatid deletions, which is typical for a normal spontaneous aberration pattern. Abnormal patterns, including significantly elevated aberration frequencies and/or presence of chromosome type exchange aberrations, were found in 13 women. The aim of the study was to evaluate a possible correlation between cytogenetic anomalies and reproductive system disorders and/or the level of radiation contamination of the territory. As a result of our estimation, it was shown that cytogenetic anomalies were in fact higher in subjects residing in territories with a higher radiation contamination; however, the difference was statistically insignificant. The frequency of cytogenetic anomalies was higher in women with gynecological disorders including hormone-dependent and inflammatory (17%) than in healthy women (7%). This difference was statistically significant for subjects with hormone-dependent diseases of the female reproductive system, but not in patients with inflammatory disorders

  7. Genomic instability in rat: Breakpoints induced by ionising radiation and interstitial telomeric-like sequences

    International Nuclear Information System (INIS)

    Camats, Nuria; Ruiz-Herrera, Aurora; Parrilla, Juan Jose; Acien, Maribel; Paya, Pilar; Giulotto, Elena; Egozcue, Josep; Garcia, Francisca; Garcia, Montserrat

    2006-01-01

    The Norwegian rat (Rattus norvegicus) is the most widely studied experimental species in biomedical research although little is known about its chromosomal structure. The characterisation of possible unstable regions of the karyotype of this species would contribute to the better understanding of its genomic architecture. The cytogenetic effects of ionising radiation have been widely used for the study of genomic instability, and the importance of interstitial telomeric-like sequences (ITSs) in instability of the genome has also been reported in previous studies in vertebrates. In order to describe the unstable chromosomal regions of R. norvegicus, the distribution of breakpoints induced by X-irradiation and ITSs in its karyotype were analysed in this work. For the X-irradiation analysis, 52 foetuses (from 14 irradiated rats) were studied, 4803 metaphases were analysed, and a total of 456 breakpoints induced by X-rays were detected, located in 114 chromosomal bands, with 25 of them significantly affected by X-irradiation (hot spots). For the analysis of ITSs, three foetuses (from three rats) were studied, 305 metaphases were analysed and 121 ITSs were detected, widely distributed in the karyotype of this species. Seventy-six percent of all hot spots analysed in this study were co-localised with ITSs

  8. Mapping of the ARIX homeodomain gene to mouse chromosome 7 and human chromosome 11q13

    Energy Technology Data Exchange (ETDEWEB)

    Johnson, K.R. [Jackson Lab., Bar Harbor, ME (United States); Smith, L.; Rhodes, J. [Oregon Health Sciences Univ., Portland, OR (United States)] [and others

    1996-05-01

    The recently described homeodomain protein ARIX is expressed specifically in noradreneric cell types of the sympathetic nervous system, brain, and adrenal medulla. ARIX interacts with regulatory elements of the genes encoding the noradrenergic biosynthetic enzymes tyrosine hydroxylase and dopamine {beta}-hydroxylase, suggesting a role for ARIX in expression of the noradrenergic phenotype. In the study described here, the mouse and human ARIX genes are mapped. Using segregation analysis of two panels of mouse backcross DNA, mouse Arix was positioned approximately 50 cM distal to the centromere of chromosome 7, near Hbb. Human ARIX was positioned through analysis of somatic cell hybrids and fluorescence in situ hybridization of human metaphase chromosomes to chromosome 7, near Hbb. Human ARIX was positioned through analysis of somatic cell hybrids and fluorescence in situ hybridization of human metaphase chromosomes to chromosome 11q13.3-q13.4. These map locations extend and further define regions of conserved synteny between mouse and human genomes and identify a new candidate gene for inherited developmental disorders linked to human 11q13.

  9. Classical and molecular cytogenetic analysis in head and neck squamous cell carcinomas

    Directory of Open Access Journals (Sweden)

    Luciana CS Veiga

    2003-01-01

    Full Text Available Head and neck carcinomas represent the sixth most frequent type of cancer in the world, and 90% are derived from squamous cells (HNSCC. In this study of 15 HNSCC cases, extensive aneuploidy was detected by G banding in most tumors. The most frequently observed numerical changes involved gain of a chromosome 22, and loss of chromosomes Y, 10, 17, and 19. The most frequent structural alteration was del(22(q13.1. As compared to G-banding, fluorescence in situ hybridization (FISH proved to be an effective technique for detecting aneuploidy. Interphase FISH with a chromosome 17 centromere probe disclosed a high frequency of monosomy for chromosome 17, in contrast with G-banding, by which clonal monosomy 17 was detected in only three of the tumors. Painting probes for chromosomes 5 and 16 were used to evaluate a selected series of HNSCC in which G-banding analysis had shown marker chromosomes. FISH analysis failed to confirm the origin of the marker chromosomes, but four out of five cases showed a significant loss of chromosomes 5. This difference between FISH and G-banding results may reflect the smaller number of metaphase analyzed as well as the criteria adopted for sorting these metaphases. Therefore results obtained solely by G-banding analysis should be considered with caution. Our data confirmed the involvement of chromosome 17 in head and neck squamous cell carcinomas.

  10. Analysis of unstable chromosome alterations frequency induced by neutron-gamma mixed field radiation

    International Nuclear Information System (INIS)

    Souza, Priscilla L.G.; Brandao, Jose Odinilson de C.; Vale, Carlos H.F.P.; Santos, Joelan A.L.; Vilela, Eudice C.; Lima, Fabiana F.; Calixto, Merilane S.; Santos, Neide

    2009-01-01

    Nowadays monitoring chromosome alterations in peripheral blood lymphocytes have been used to access the radiation absorbed dose in individuals exposed accidental or occupationally to gamma radiation. However there are not many studies based on the effects of mixed field neutron-gamma. The radiobiology of neutrons has great importance because in nuclear factories worldwide there are several hundred thousand individuals monitored as potentially receiving doses of neutron. In this paper it was observed the frequencies of unstable chromosome alterations induced by a gamma-neutron mixed field. Blood was obtained from one healthy donor and exposed to mixed field neutron-gamma sources 241 AmBe (20 Ci) at the Neutron Calibration Laboratory (NCL-CRCN/NE-PE-Brazil). The chromosomes were observed at metaphase, following colcemid accumulation and 1000 well-spread metaphases were analyzed for the presence of chromosome alterations by two experienced scorers. The results suggest that there is the possibility of a directly proportional relationship between absorbed dose of neutron-gamma mixed field radiation and the frequency of unstable chromosome alterations analyzed in this paper. (author)

  11. Kinetochore–microtubule attachment throughout mitosis potentiated by the elongated stalk of the kinetochore kinesin CENP-E

    Science.gov (United States)

    Vitre, Benjamin; Gudimchuk, Nikita; Borda, Ranier; Kim, Yumi; Heuser, John E.; Cleveland, Don W.; Grishchuk, Ekaterina L.

    2014-01-01

    Centromere protein E (CENP-E) is a highly elongated kinesin that transports pole-proximal chromosomes during congression in prometaphase. During metaphase, it facilitates kinetochore–microtubule end-on attachment required to achieve and maintain chromosome alignment. In vitro CENP-E can walk processively along microtubule tracks and follow both growing and shrinking microtubule plus ends. Neither the CENP-E–dependent transport along microtubules nor its tip-tracking activity requires the unusually long coiled-coil stalk of CENP-E. The biological role for the CENP-E stalk has now been identified through creation of “Bonsai” CENP-E with significantly shortened stalk but wild-type motor and tail domains. We demonstrate that Bonsai CENP-E fails to bind microtubules in vitro unless a cargo is contemporaneously bound via its C-terminal tail. In contrast, both full-length and truncated CENP-E that has no stalk and tail exhibit robust motility with and without cargo binding, highlighting the importance of CENP-E stalk for its activity. Correspondingly, kinetochore attachment to microtubule ends is shown to be disrupted in cells whose CENP-E has a shortened stalk, thereby producing chromosome misalignment in metaphase and lagging chromosomes during anaphase. Together these findings establish an unexpected role of CENP-E elongated stalk in ensuring stability of kinetochore–microtubule attachments during chromosome congression and segregation. PMID:24920822

  12. Biological radiation dose estimation by chromosomal aberrations analysis in human peripheral blood (dose-effect curve)

    International Nuclear Information System (INIS)

    Al-Achkar, W.

    2001-09-01

    In order to draw a dose-effect curve, experimentally gamma ray induced chromosomal aberrations in human peripheral lymphocytes from eight healthy people were studied. Samples from 4 males and 4 females were irradiated in tubes with 0.15, 0.25, 0.5, 1, 1.5, 2, 2.5 gray of gamma ray (Co 60 at dose rate 0.3 Gy/min). Irradiated and control samples were incubated in 37 centigrade for 48 hours cell cultures. Cell cultures then were stopped and metaphases spread, Giemsa stained to score the induced chromosomal aberrations. Chromosomal aberrations from 67888 metaphases were scored. Curves from the total number of dicentrics, dicentrics + rings and total numbers of breaks in cell for each individual or for all people were drawn. An increase of all chromosomal aberrations types with the elevation of the doses was observed. The yield of chromosome aberrations is related to the dose used. These curves give a quick useful estimation of the accidentally radiation exposure. (author)

  13. The spatio-temporal dynamics of PKA activity profile during mitosis and its correlation to chromosome segregation

    Science.gov (United States)

    Vandame, Pauline; Spriet, Corentin; Trinel, Dave; Gelaude, Armance; Caillau, Katia; Bompard, Coralie; Biondi, Emanuele; Bodart, Jean-François

    2014-01-01

    The cyclic adenosine monophosphate dependent kinase protein (PKA) controls a variety of cellular processes including cell cycle regulation. Here, we took advantages of genetically encoded FRET-based biosensors, using an AKAR-derived biosensor to characterize PKA activity during mitosis in living HeLa cells using a single-cell approach. We measured PKA activity changes during mitosis. HeLa cells exhibit a substantial increase during mitosis, which ends with telophase. An AKAREV T>A inactive form of the biosensor and H89 inhibitor were used to ascertain for the specificity of the PKA activity measured. On a spatial point of view, high levels of activity near to chromosomal plate during metaphase and anaphase were detected. By using the PKA inhibitor H89, we assessed the role of PKA in the maintenance of a proper division phenotype. While this treatment in our hands did not impaired cell cycle progression in a drastic manner, inhibition of PKA leads to a dramatic increase in chromososme misalignement on the spindle during metaphase that could result in aneuploidies. Our study emphasizes the insights that can be gained with genetically encoded FRET-based biosensors, which enable to overcome the shortcomings of classical methologies and unveil in vivo PKA spatiotemporal profiles in HeLa cells. PMID:25485503

  14. Nucleolar-persistence phenomenon during spermatogenesis in genus Meccus (Hemiptera, Triatominae).

    Science.gov (United States)

    Madeira, F F; Lima, A A C; Rosa, J A; Azeredo-Oliveira, M T V; Alevi, K C C

    2016-03-28

    The Triatominae subfamily consists of 150 species in 18 genera, grouped into six tribes. In cytogenetics, triatomines are important biological models because they have holocentric chromosomes and nucleolar persistence in meiosis. The phenomenon of nucleolar persistence has been described for 23 species of triatomine in three genera: Triatoma, Rhodnius, and Panstrongylus. However, new species and genera should be analyzed to assess whether nucleolar persistence is a peculiarity of Triatominae. Thus, this study aimed to analyze nucleolar behavior during spermatogenesis of Meccus pallidipennis and M. longipennis, focusing on the nucleolar-persistence phenomenon. Through the analysis of spermatogenesis, more specifically of meiotic metaphase, we observed the phenomenon of nucleolar persistence in M. pallidipennis and M. longipennis, represented by remnants of nucleolar material in metaphase. Thus, although nucleologenesis of new species, and, especially, new genera, should be analyzed, this study confirms for the first time the phenomenon of nucleolar persistence in the genus Meccus. Therefore, we emphasize the importance of new studies in this area in order to assess whether this phenomenon is truly a synapomorphy of these hematophagous insects.

  15. Radioactive probes for human gene localisation by in situ hybridisation

    International Nuclear Information System (INIS)

    Fennell, S.J.

    1980-07-01

    Radioactive probes of high specific activity have been used for human gene localisation on metaphase chromosome preparations. Human 5S ribosomal RNA was used as a model system, as a probe for the localisation of human 5S ribosomal genes. 125 I-labelled mouse 5S ribosomal RNA was used to study the 5S ribosomal gene content and arrangement in families with translocations on the long arm of chromosome 1 close to or containing the 5S ribosomal RNA locus, by in situ hybridisation to human metaphase chromosomes from peripheral blood cultures. This confirmed the chromosomal assignment of 5S ribosomal genes to 1q 42-43. In situ hybridisation probes were also prepared from recombinant plasmids containing Xenopus laevis oocyte 5S or 28S/18S gene sequences to give [ 3 H]-labelled cRNA and [ 3 H]-labelled nick-translated plasmid DNA. Studies on the kinetics of hybridisation of plasmid probes with and without ribosomal gene sequences questioned the role of plasmid DNA for amplification of signal during gene localisation. Gene localisation was obtained with nick-translated plasmid DNA containing the 28S/18S ribosomal DNA insert after short exposure times, but poor results were obtained using a [ 3 H]-labelled cRNA probe transcribed from the plasmid with the 5S gene insert. (author)

  16. The spatio-temporal dynamics of PKA activity profile during mitosis and its correlation to chromosome segregation.

    Science.gov (United States)

    Vandame, Pauline; Spriet, Corentin; Trinel, Dave; Gelaude, Armance; Caillau, Katia; Bompard, Coralie; Biondi, Emanuele; Bodart, Jean-François

    2014-01-01

    The cyclic adenosine monophosphate dependent kinase protein (PKA) controls a variety of cellular processes including cell cycle regulation. Here, we took advantages of genetically encoded FRET-based biosensors, using an AKAR-derived biosensor to characterize PKA activity during mitosis in living HeLa cells using a single-cell approach. We measured PKA activity changes during mitosis. HeLa cells exhibit a substantial increase during mitosis, which ends with telophase. An AKAREV T>A inactive form of the biosensor and H89 inhibitor were used to ascertain for the specificity of the PKA activity measured. On a spatial point of view, high levels of activity near to chromosomal plate during metaphase and anaphase were detected. By using the PKA inhibitor H89, we assessed the role of PKA in the maintenance of a proper division phenotype. While this treatment in our hands did not impaired cell cycle progression in a drastic manner, inhibition of PKA leads to a dramatic increase in chromososme misalignement on the spindle during metaphase that could result in aneuploidies. Our study emphasizes the insights that can be gained with genetically encoded FRET-based biosensors, which enable to overcome the shortcomings of classical methologies and unveil in vivo PKA spatiotemporal profiles in HeLa cells.

  17. Chromosome Damage and Cell Proliferation Rates in In Vitro Irradiated Whole Blood as Markers of Late Radiation Toxicity After Radiation Therapy to the Prostate

    Energy Technology Data Exchange (ETDEWEB)

    Beaton, Lindsay A., E-mail: Lindsay.Beaton@hc-sc.gc.ca [Environmental and Radiation Health Sciences Directorate, Health Canada, Ottawa, ON (Canada); Ferrarotto, Catherine; Marro, Leonora [Environmental and Radiation Health Sciences Directorate, Health Canada, Ottawa, ON (Canada); Samiee, Sara; Malone, Shawn; Grimes, Scott; Malone, Kyle [The Ottawa Hospital, Ottawa Hospital Research Institute, University of Ottawa, 501 Smyth Rd, Ottawa, ON (Canada); Wilkins, Ruth C. [Environmental and Radiation Health Sciences Directorate, Health Canada, Ottawa, ON (Canada)

    2013-04-01

    Purpose: In vitro irradiated blood samples from prostate cancer patients showing late normal tissue damage were examined for lymphocyte response by measuring chromosomal aberrations and proliferation rate. Methods and Materials: Patients were selected from a randomized trial evaluating the optimal timing of dose-escalated radiation and short-course androgen deprivation therapy. Of 438 patients, 3% experienced grade 3 late radiation proctitis and were considered to be radiosensitive. Blood samples were taken from 10 of these patients along with 20 matched samples from patients with grade 0 proctitis. The samples were irradiated at 6 Gy and, along with control samples, were analyzed for dicentric chromosomes and excess fragments per cell. Cells in first and second metaphase were also enumerated to determine the lymphocyte proliferation rate. Results: At 6 Gy, there were statistically significant differences between the radiosensitive and control cohorts for 3 endpoints: the mean number of dicentric chromosomes per cell (3.26 ± 0.31, 2.91 ± 0.32; P=.0258), the mean number of excess fragments per cell (2.27 ± 0.23, 1.43 ± 0.37; P<.0001), and the proportion of cells in second metaphase (0.27 ± 0.10, 0.46 ± 0.09; P=.0007). Conclusions: These results may be a valuable indicator for identifying radiosensitive patients and for tailoring radiation therapy.

  18. Noninvasive three-dimensional live imaging methodology for the spindles at meiosis and mitosis

    Science.gov (United States)

    Zheng, Jing-gao; Huo, Tiancheng; Tian, Ning; Chen, Tianyuan; Wang, Chengming; Zhang, Ning; Zhao, Fengying; Lu, Danyu; Chen, Dieyan; Ma, Wanyun; Sun, Jia-lin; Xue, Ping

    2013-05-01

    The spindle plays a crucial role in normal chromosome alignment and segregation during meiosis and mitosis. Studying spindles in living cells noninvasively is of great value in assisted reproduction technology (ART). Here, we present a novel spindle imaging methodology, full-field optical coherence tomography (FF-OCT). Without any dye labeling and fixation, we demonstrate the first successful application of FF-OCT to noninvasive three-dimensional (3-D) live imaging of the meiotic spindles within the mouse living oocytes at metaphase II as well as the mitotic spindles in the living zygotes at metaphase and telophase. By post-processing of the 3-D dataset obtained with FF-OCT, the important morphological and spatial parameters of the spindles, such as short and long axes, spatial localization, and the angle of meiotic spindle deviation from the first polar body in the oocyte were precisely measured with the spatial resolution of 0.7 μm. Our results reveal the potential of FF-OCT as an imaging tool capable of noninvasive 3-D live morphological analysis for spindles, which might be useful to ART related procedures and many other spindle related studies.

  19. Identification of a YAC spanning the translocation breakpoints in uterine leiomyomata, pulmonary chondroid hamartoma, and lipoma: Physical mapping of the 12q14-q15 breakpoint region in uterine leiomyomata

    Energy Technology Data Exchange (ETDEWEB)

    Fejzo, M.S. [Harvard Medical School, Boston, MA (United States); Yoon, S.J.; Kucherlapati, R.S. [Albert Einstein College of Medicine, Bronx, NY (United States)] [and others

    1995-03-20

    Uterine leiomyomata are the most common tumors in women and can cause abnormal uterine bleeding, pelvic pain, and infertility. Approximately 200,000 hysterectomies are performed annually in the U.S. to relieve patients of the medical sequelae of these benign neoplasms. Our efforts have focused on cloning the t(12;14)(q14-q15;q23-q24) breakpoint in uterine leiomyoma to further our understanding of the biology of these tumors. Thirty-nine YACs and six cosmids mapping to 12q14-q15 have been mapped by fluorescence in situ hybridization to tumor metaphase chromosomes containing a t(12;14). One YAC spanned the translocation breakpoint and was mapped to tumor metaphases from a pulmonary chondroid hamartoma containing a t(12;14)(q14-q15;q23-q24) and a lipoma containing a t(12;15)(q15;q24); this YAC also spanned the breakpoint in these two tumors, suggesting that the same gene on chromosome 12 may be involved in the pathobiology of these distinct benign neoplasms. 41 refs., 2 figs., 1 tab.

  20. Histone H2AX phosphorylation is associated with most meiotic events in grasshopper.

    Science.gov (United States)

    Cabrero, J; Teruel, M; Carmona, F D; Camacho, J P M

    2007-01-01

    It is widely accepted that the H2AX histone in its phosphorylated form (gamma-H2AX) is related to the repair of DNA double-strand breaks (DSBs). In several organisms, gamma-H2AX presence has been demonstrated in meiotic processes such as recombination and sex chromosome inactivation during prophase I (from leptotene to pachytene). To test whether gamma-H2AX is present beyond pachytene, we have analysed the complete sequence of changes in H2AX phosphorylation during meiosis in grasshopper, a model organism for meiotic studies at the cytological level. We show the presence of phosphorylated H2AX during most of meiosis, with the exception only of diplotene and the end of each meiotic division. During the first meiotic division, gamma-H2AX is associated with i) recombination, as deduced from its presence in leptotene-zygotene over all chromosome length, ii) X chromosome inactivation, since at pachytene gamma-H2AX is present in the X chromosome only, and iii) chromosome segregation, as deduced from gamma-H2AX presence in centromere regions at first metaphase-anaphase. During second meiotic division, gamma-H2AX was very abundant at most chromosome lengths from metaphase to telophase, suggesting its possible association with the maintenance of chromosome condensation and segregation. Copyright 2007 S. Karger AG, Basel.

  1. Granulocytes with segmented nucleus retain normal chromosomes 17 in Philadelphia chromosome-positive chronic myeloid leukemia with i(17q) and pseudo-Pelger anomaly. A case report studied with fluorescence in situ hybridization.

    Science.gov (United States)

    Fugazza, G; Bruzzone, R; Puppo, L; Sessarego, M

    1996-09-01

    Previous reports suggested a correlation between the deletion of the terminal region of the short arm of a chromosome 17 and the appearance of dysgranulopoiesis in myeloproliferative disorders. Using the dual-color fluorescence in situ hybridization technique we analyzed the bone marrow and peripheral blood cells of a Philadelphia chromosome-positive chronic myeloid leukemia (CML) patient showing at the onset of transformation into blastic crisis both metaphases with the i(17q) as well as granulocytes without nuclear segmentation. This phenomenon is defined as pseudo-Pelger-Huët anomaly. Using two probes, one specific for 17p and one for 17q, we determined the presence or absence of the i(17q) in both metaphase and interphase cells. Moreover, we observed that all cells with a polysegmented nucleus typical of mature granulocytes did not have i(17q) but had two normal chromosomes 17. This observation confirmed the correlation between 17p deletion and the appearance of pseudo-Pelger anomaly. This finding may also be useful from a clinical point of view: the appearance of pseudo-Pelger cells in CML indicates that 17p deletion actually occurred. This event implies a negative prognosis.

  2. Philadelphia chromosome detection in chronic myeloid leukemia: Utility of phytohemagglutinin-stimulated peripheral blood culture

    Directory of Open Access Journals (Sweden)

    Man Updesh Singh Sachdeva

    2012-01-01

    Full Text Available Background: The conventional cytogenetic approach to demonstrate Philadelphia (Ph chromosome at times does not yield enough number of metaphases or are of suboptimal quality. Further, the rapid molecular tests have completely pushed this simple technique into disrepute. Aims: This study aimed to evaluate usefulness of phytohemagglutinin (PHA-stimulated peripheral blood culture for detection of Ph chromosome in chronic myeloid leukemia (CML patients. Materials and Methods: Fifty-six patients, including 11 newly diagnosed cases of CML and 45 patients of CML on imatinib therapy showing the presence of Ph chromosome in unstimulated samples, were included in the study. Cytogenetic analysis was done on unstimulated samples, i.e. bone marrow aspirate, 24- and 48-h peripheral blood culture, and compared with PHA-stimulated 72-h peripheral blood culture. Results: The preparations from PHA-stimulated peripheral blood culture samples in all 56 patients yielded high number of good-quality metaphases. All the 11 (100% newly diagnosed patients and 39/45 (87% of the patients on imatinib therapy showed the presence of Ph chromosome in PHA-stimulated samples. Addition of PHA-stimulated 72-h peripheral blood culture preparation can be of use for increasing the diagnostic yield in cases of CML with suboptimal results on conventional cytogenetics from bone marrow aspirate sample.

  3. X-ray induced polyploidization in the male germline cells of Poekilocerus pictus (acrididoidea : orthopta)

    International Nuclear Information System (INIS)

    Gururaj, M.E.; Rajasekarasetty, M.R.

    1977-01-01

    After the irradiation of male germline cells of Poekilocerus pictus with 20r, 40r, 80r, 120r doses of X-rays, both first and second meiotic polyploid cells were recovered. While various degrees of polyploidy were encountered in first meiotic cells, second meiotic polyploid cells, second meitoic polyploid cells contained diploid number of half bivalents only. The former never progressed beyond leptotene and showed symptoms of degeneration. Among the latter, a few cells showed either emainingative tendencies like uncoiling and stickiness or failure of cellsted meiosis successfully. It has been shown that the dicentric bridges and/or laggards in anaphase-I interfere with the elongation and regression of the spindle, thereby giving rise to metaphase-II polyploid cells through restitution. The possible role of fragmentation of chromosomes in decreasing the incidence of metaphase-II polyploid cells at higher doses of irradiation and the causes for the differential fate of the first and second meiotic polyploid cells have been discussed. (author)

  4. Analysis of meiotic behavior in Cordia ecalyculata Vell. (Boraginaceae

    Directory of Open Access Journals (Sweden)

    Damião do Nascimento

    2014-12-01

    Full Text Available Cordia ecalyculata belongs to the Boraginaceae family, and is commonly known as buggy coffee. It is indicated for medicinal use as a tonic, diuretic, anti-inlammatory and appetite suppressant. Young inlorescences of six individuals were collected and ixed in a mixture of ethanol and acetic acid (3:1 for 24 hours. The slides were prepared by crushing and staining tissue with 1% acetic carmine. During microsporogenesis some irregularities were observed, mostly frequently related to irregular chromosome segregation. Irregularities included: precocious migration to poles in metaphase I and II, disoriented bivalent chromosomes at metaphase I and II, laggard chromosomes in anaphase I and II, and micronuclei formation. We also observed irregular spindle organization in meiosis II, leading to ‘T’ and ‘V’ shaped spindle conigurations. In the V-shaped coniguration, two nearby nuclei fused, forming triads instead of tetrads; this lead to formation of 2n microspore at the end of meiosis. However, pollen grain viability was not compromised, as pollen grain viability varied between 95.42% and 100%.

  5. Phosphatase-regulated recruitment of the spindle- and kinetochore-associated (Ska complex to kinetochores

    Directory of Open Access Journals (Sweden)

    Sushama Sivakumar

    2017-11-01

    Full Text Available Kinetochores move chromosomes on dynamic spindle microtubules and regulate signaling of the spindle checkpoint. The spindle- and kinetochore-associated (Ska complex, a hexamer composed of two copies of Ska1, Ska2 and Ska3, has been implicated in both roles. Phosphorylation of kinetochore components by the well-studied mitotic kinases Cdk1, Aurora B, Plk1, Mps1, and Bub1 regulate chromosome movement and checkpoint signaling. Roles for the opposing phosphatases are more poorly defined. Recently, we showed that the C terminus of Ska1 recruits protein phosphatase 1 (PP1 to kinetochores. Here we show that PP1 and protein phosphatase 2A (PP2A both promote accumulation of Ska at kinetochores. Depletion of PP1 or PP2A by siRNA reduces Ska binding at kinetochores, impairs alignment of chromosomes to the spindle midplane, and causes metaphase delay or arrest, phenotypes that are also seen after depletion of Ska. Artificial tethering of PP1 to the outer kinetochore protein Nuf2 promotes Ska recruitment to kinetochores, and it reduces but does not fully rescue chromosome alignment and metaphase arrest defects seen after Ska depletion. We propose that Ska has multiple functions in promoting mitotic progression and that kinetochore-associated phosphatases function in a positive feedback cycle to reinforce Ska complex accumulation at kinetochores.

  6. Cytogenetic data on the agro-predatory ant Megalomyrmex incisus Smith, 1947 and its host, Mycetophylax conformis (Mayr, 1884) (Hymenoptera, Formicidae)

    Science.gov (United States)

    Cardoso, Danon Clemes; Pereira, Tássia Tatiane Pontes; Cordeiro, Alessandro Lick; Cristiano, Maykon Passos

    2017-01-01

    Abstract We provide the first karyotype description of the agro-predatory ant species Megalomyrmex incisus Smith, 1947 (Myrmicinae, Formicidae), and chromosome counts of its host Mycetophylax conformis (Mayr, 1884) (Myrmicinae, Formicidae) from geographically distinct populations. Colonies of both species were sampled from coastal areas of Ilhéus, Bahia, Brazil, and transferred to the laboratory. Metaphase spreads were prepared from the cerebral ganglia of defecated larvae. The slides were examined and pictures of the best metaphases were taken. The chromosome number for Megalomyrmex incisus was 2n=50 and n=25. The karyotype of this species consists of 20 metacentric and 5 submetacentric pairs. Thus, the karyotype formula of the diploid set was 2K=40M + 10SM and a fundamental number FN=100. The host species Mycetophylax conformis has 2n=30 and the karyotype consisting of 11 metacentric and 4 submetacentric pairs. The karyotype formula was 2K=22M + 8SM, and a fundamental number FN=60. Megalomyrmex incisus showed a slightly higher chromosome number, placed at the marginal range of the known distribution of haploid karyotypes of the Myrmicinae. The chromosome number and chromosomal morphology of Mycetophylax conformis corresponded to those of previously studied populations, suggesting its karyotype stability. PMID:28919948

  7. Cumulus cells gene expression profiling in terms of oocyte maturity in controlled ovarian hyperstimulation using GnRH agonist or GnRH antagonist.

    Science.gov (United States)

    Devjak, Rok; Fon Tacer, Klementina; Juvan, Peter; Virant Klun, Irma; Rozman, Damjana; Vrtačnik Bokal, Eda

    2012-01-01

    In in vitro fertilization (IVF) cycles controlled ovarian hyperstimulation (COH) is established by gonadotropins in combination with gonadotropin-releasing hormone (GnRH) agonists or antagonists, to prevent premature luteinizing hormone (LH) surge. The aim of our study was to improve the understanding of gene expression profile of cumulus cells (CC) in terms of ovarian stimulation protocol and oocyte maturity. We applied Affymetrix gene expression profiling in CC of oocytes at different maturation stages using either GnRH agonists or GnRH antagonists. Two analyses were performed: the first involved CC of immature metaphase I (MI) and mature metaphase II (MII) oocytes where 359 genes were differentially expressed, and the second involved the two GnRH analogues where no differentially expressed genes were observed at the entire transcriptome level. A further analysis of 359 differentially genes was performed, focusing on anti-Müllerian hormone receptor 2 (AMHR2), follicle stimulating hormone receptor (FSHR), vascular endothelial growth factor C (VEGFC) and serine protease inhibitor E2 (SERPINE2). Among other differentially expressed genes we observed a marked number of new genes connected to cell adhesion and neurotransmitters such as dopamine, glycine and γ-Aminobutyric acid (GABA). No differential expression in CC between the two GnRH analogues supports the findings of clinical studies where no significant difference in live birth rates between both GnRH analogues has been proven.

  8. Effect of low 60Co dose rates on sister chromatid exchange incidence in the benthic worm. Neanthes arenaceodentata

    International Nuclear Information System (INIS)

    Harrison, F.L.; Rice, D.W. Jr.

    1981-01-01

    The usefulness of sister chromatid exchange (SCE) induction as a measure of low-level radiation effect was examined in a benthic marine worm, Neanthes arenaceodentata. Larvae were exposed to 60 Co radiation for 12 to 24 h at total doses ranging from 0.5 to 309 R and at dose rates from 0.04 to 13 R/h. Animals exposed at intermediate dose rates (0.5, 0.6, 1.25, 2.0, and 2.5 R/h) had SCE frequencies per chromosome about twice that of those receiving no radiation (controls), whereas those exposed at the higher dose rates (7.0 and 13 R/h) had SCE frequencies lower than the controls. Animals exposed at the lower dose rates (0.04 and 0.1 R/h) had lower SCE frequencies than those exposed at intermediate dose rates (and higher SCE frequencies than controls). The length of chromosome pair number one differed among metaphase spreads and was used as an index of chromosome condensation in a given metaphase. Because there is a possibility that chromosome morphology may affect the ability to resolve SCEs, morphology will be monitored in future studies. A preliminary experiment was performed to assess the effects of 2.2 and 11.5 R/h for 24 h on growth and development. Larvae observed at 6 and 17 d after irradiation did not have significantly different numbers of abnormal larvae or survival rates

  9. The importance of using fluorescence in situ hybridization for the diagnosis of Smith-Magenis syndrome

    Energy Technology Data Exchange (ETDEWEB)

    Juyal, R.C.; Greenberg, F.; Lupski, J.R. [Baylor College of Medicine, Houston, TX (United States)] [and others

    1994-09-01

    Smith-Magenis syndrome (SMS) is a clinically recognizable multiple congenital anomaly/mental retardation syndrome associated with deletion of chromosome 17p11.2. Quality metaphase preparations are required for unambiguous detection of the deletion. We and others have reported cases of SMS due to mosaicism for del(17)(p11.2). Examination of peripheral blood lymphocyte cultures of a patient with the SMS phenotype at 850 band level of resolution revealed a low level mosaicism (11%) for the deletion. Examination of fibroblasts at relatively low resolution revealed the deletion in all cells. In a second study, we reported molecular evidence for mosaicism in the unaffected mother of an SMS patient who demonstrated mosaicism (55%) for the deletion at a resolution level of < 500 bands. We now report a different SMS patient who was initially diagnosed as mosaic del(17)(p11.2) in two different cytogenetic laboratories. A third blinded cytogenetic study yielded a questionable diagnosis. Fluorescence in situ hybridization (FISH) conducted in two different laboratories with two different markers shown to be within the deletion region and a control marker from chromosome 17 demonstrated a deletion in 20/20 and 25/25 metaphases scored, respectively. It appears the latter patient may harbor a very small deletion and that FISH is a more reliable test for the Smith-Magenis deletion. Furthermore, FISH should be used to confirm or refute mosaicism seen in routine cytogenetics studies.

  10. Comparison of three in vitro culture systems for maturation of early preantral mouse ovarian follicles.

    Science.gov (United States)

    Mousset-Simeón, Nathalie; Jouannet, Pierre; Le Cointre, Laëtitia; Coussieu, Christiane; Poirot, Catherine

    2005-05-01

    The aim of this study was to compare three different culture systems for in vitro follicular growth and oocyte maturation in ovarian follicles of mice in order to assess the technique with the optimal growth and improved rate of meiotic maturation. The three systems tested were culture under oil, on a hydrophobic membrane and on agar respectively. Early preantral follicles were cultured for 12 days in alpha-MEM GlutaMAX medium. Follicular growth, oocyte meiotic maturation, oocyte extrusion, atresia and estradiol production were analysed. Follicular development showed two phases in the three systems, with slow growth before day 5 and subsequent acceleration. The percentage of follicles transferred into oocyte maturation medium was significantly higher after culture under oil. The proportion of oocytes that achieved nuclear maturation (metaphase II) was higher when follicles were cultured under oil or on a hydrophobic membrane than on agar. Our results support the use of culture under oil for in vitro follicular growth from the early preantral stage in order to obtain metaphase II oocytes. Fertilization ability of these oocytes and the capacity to obtain healthy mice in a reproducible manner warrants further investigation.

  11. Molecular cytogenetic characterization of inv dup del(8p) in a fetus associated with ventriculomegaly, hypoplastic left heart, polyhydramnios and intestinal obstruction.

    Science.gov (United States)

    Chen, Chih-Ping; Ko, Tsang-Ming; Huang, Wen-Chu; Chern, Schu-Rern; Wu, Peih-Shan; Chen, Yen-Ni; Chen, Shin-Wen; Lee, Chen-Chi; Pan, Chen-Wen; Yang, Chien-Wen; Wang, Wayseen

    2016-06-01

    To present molecular cytogenetic characterization of inv dup del(8p) in a fetus with congenital malformations. A 19-year-old, primigravid woman underwent cord blood sampling at 31 weeks of gestation because of prenatal ultrasound findings of polyhydramnios, intestinal obstruction, right ventriculomegaly, and hypoplastic left heart. Preterm precipitous labor and delivery occurred at 32 weeks of gestation. Array comparative genomic hybridization (aCGH), conventional cytogenetic analysis and metaphase fluorescence in situ hybridization (FISH) were applied on cord blood lymphocytes. aCGH was also applied on the umbilical cord. Conventional cytogenetic analysis was applied on parental bloods. aCGH detected an 11.35 Mb deletion in 8p23.3-p23.1 encompassing SOX7 and GATA4, and a 31.99 Mb duplication in 8p23.1-p11.1 in the fetus. Metaphase FISH confirmed inv dup del(8p). The fetus had a karyotype of 46,XX,der(8)del(8)(p23.1) inv dup(8) (p11.1p23.1). Parental karyotypes were normal. A malformed fetus was delivered with facial dysmorphism. Fetuses with inv dup del(8p) may present central nervous system (CNS) abnormality and congenital heart defect on prenatal ultrasound. Prenatal diagnosis of concomitant CNS and cardiac abnormalities should include a differential diagnosis of chromosome 8p inverted duplication deletion syndrome. Copyright © 2016. Published by Elsevier B.V.

  12. Induction of chromosomal aberrations in mouse zygotes by acrylamide treatment of male germ cells and their correlation with dominant lethality and heritable translocations

    Energy Technology Data Exchange (ETDEWEB)

    Marchetti, F.; Lowe, X.; Wyrobek, A.J. [Lawrence Livermore National Lab., CA (United States); Bishop, J. [National Institute of Environmental Health Sciences, Research Triangle Park, NC (United States)

    1997-12-31

    The objectives of this research were: (1) to investigate the time course of the cytogenetic defects induced by acrylamide (AA) treatment (5 x 50 mg/kg) of male germ cells in first-cleavage zygote metaphases using PAINT/DAPI analysis, and (2) to characterize the correlation between chromosomal aberrations at first cleavage, dominant lethality, and heritable translocations. PAINT/DAPI analysis employs multicolor fluorescence in situ hybridization painting plus DAPI staining to detect both stable and unstable chromosomal aberrations at first-cleavage metaphase of the zygote. High levels of chromosomally defective zygotes were detected after mating at all postmeiotic stages (20-190-fold, P < 0.001). Early spermatozoa (6.5 d post-treatment) were the most sensitive, with 76% of the zygotes carrying cytogenetic defects. A significant 10-fold increase was also detected 27.5 d post-treatment, indicating that AA had a cytogenetic effect on meiotic stages. PAINT/DAPI analysis revealed that: (1) AA-induced chromosomal breaks occurred at random, and (2) the frequencies of symmetrical and asymmetrical exchanges were similar at all mating days, except 9.5 d after AA treatment, where significantly (P < 0.02) more asymmetrical aberrations were found. 33 refs., 5 figs., 4 tabs.

  13. In vivo wound healing and antiulcer properties of white sweet potato (Ipomoea batatas)

    Science.gov (United States)

    Hermes, Daniele; Dudek, Débora N.; Maria, Mariana D.; Horta, Lívia P.; Lima, Eliete N.; de Fátima, Ângelo; Sanches, Andréia C.C.; Modolo, Luzia V.

    2012-01-01

    The potential of tuber flour of Ipomoea batatas (L.) Lam. cv. Brazlândia Branca (white sweet potato) as wound healing and antiulcerogenic agent was investigated in vivo in animal model. Excision on the back of Wistar rats was performed to induce wounds that were topically treated with Beeler’s base containing tuber flour of white sweet potato at 2.5%. Number of cells undergoing metaphase and the degree of tissue re-epithelialization were investigated 4, 7 and 10 days post-treatment. The protective effect of aqueous suspension of tuber flour (75 and 100 mg/kg animal weight) on gastric mucosa of Wistar rats was also studied by using the ethanol-induced ulceration model. Ointment based on white sweet potato at 2.5% effectively triggered the healing of cutaneous wound as attested by the increased number of cells undergoing metaphase and tissue re-epithelialization regardless the time of wound treatment. Tuber flour potentially prevented ethanol-induced gastric ulceration by suppressing edema formation and partly protecting gastric mucosa wrinkles. Crude extracts also exhibited potential as free radical scavengers. The results from animal model experiments indicate the potential of tuber flour of white sweet potato to heal wounds. PMID:25685447

  14. Relationship Between Genotype Variants Follicle-stimulating Hormone Receptor Gene Polymorphisms (FSHR) and Morphology of Oocytes Prior to ICSI Procedures

    Science.gov (United States)

    Gashi, Zafer; Elezaj, Shkelzen; Zeqiraj, Afrim; Grabanica, Driton; Shabani, Isak; Gruda, Bujar; Gashi, Fitore

    2016-01-01

    Introduction: This study investigated association of Asn680Ser FSHR polymorphism with the ovarian response in 104 women of Albanian ethnic population enrolled in ICSI program. The reason of infertility in all cases has been identified as male factor. Methods: Analysis of the Asn680Ser polymorphism was performed using TaqMan® SNP Genotyping Assay. Clinical and endocrinologic parameters were analyzed based on the genotype, age, BMI, oocyte yield, number of transferred embryos and pregnancy rate. Results: The frequencies of the Asn680 Ser genotype variants were as follows: Asn/Asn 22.1%, Asn/Ser 47.1%, and Ser/Ser 30.8%, respectively. BMI was significantly higher in the Ser/Ser group as compared to those from the Asn/Ser or the Asn/Asn group (p= 0.0010). The genotype variants Ser/Ser indicates a higher rate of oocyte retrieval (25.9%) in the immature form, metaphase I (MI) as opposed to the other two groups (Asn/Asn 23.7 % vs. Asn/Ser 21.9%), which was statistically significant (p = 0.3020). Conclusions: FSH receptor polymorphism is associated with different ovarian response to controlled ovarian stimulation (COS), but is not an important factor in increasing the degree of pregnancy. Polymorphisms of the FSH receptor is associated with normal morphology and genetic maturation (metaphase II) oocytes in dependence of genotypic variation polymorphisms. PMID:27994298

  15. Chromosome painting analysis of X-ray-induced aberrations in human lymphocytes in vitro

    International Nuclear Information System (INIS)

    Matsuoka, A.; Hayashi, M.; Yamazaki, N.; Sofuni, T.

    1994-01-01

    Chromosomal rearrangements in human lymphocytes induced by X-rays (0, 0.5, 1.0 and 2.0 Gray) were analyzed using chromosome painting. DNA probes for human chromosomes 1, 3 or 4 alone, and a combination of 1 and 4, were used for analysis. The frequency of cells with rearrangements, i.e. reciprocal translocations, dicentrics, insertions, tricentrics and fragments, involving chromosome 4 increased with dose in both 48 and 72 h cultures. The number of translocations per cell also increased with dose at 48 and 72 h. Dicentrics increased with dose in 48 h but not in 72 h cultures. The estimated genomic frequency of aberrations per cell was comparable with results in banded cells. No difference was shown on the detection efficiency of chromosome rearrangements among the various DNA probes used. Since this technique does not necessarily require well-spread metaphases for analysis, it is possible to increase the number of analyzable metaphases compared with the banding technique. Chromosome painting is a simpler, more objective and practical method for detecting chromosome rearrangements than conventional banding analyses. (Author)

  16. [Cytogenetic features of teenage girls with secondary amenorrhea].

    Science.gov (United States)

    Nachetova, T A; Nefidova, V E

    2014-11-01

    Some features of the chromosome apparatus status were studied in 25 adolescent girls, aged 14-18, with secondary amenorrhea and in 29 girls of the same age with a regular menstrual cycle. Materials for cytogenetic analysis were preparations of chromosomes at the stage of metaphase obtained from the culture of the peripheral blood lymphocytes. The technique of the culture preparation was carried out according to the standard method. 2225 metaphase plates were analyzed in girls with secondary amenorrhea, and 2603 plates were tested in their healthy age-mates. An increased total level of chromosomal aberrations and a rise in the frequency of disorders in the chromatid, chromosome and genome types of peripheral blood lymphocytes have been registered in the examined persons as compared with their healthy age-mates. We have shown, that polyploid cell registered in 15 times oftener in adolescent girls with SA as compared with healthy girls. It can be assumed that some marked changes in the frequency of chromosomal aberrations in patients with secondary amenorrhea and in their healthy age-mates may arise both as a result of exposure to the multiple environmental factors and disorders of rather complicated processes of DNA damages reparation.

  17. Dose - Response Curves for Dicentrics and PCC Rings: Preparedness for Radiological Emergency in Thailand

    International Nuclear Information System (INIS)

    Rungsimaphorn, B.; Rerkamnuaychoke, B.; Sudprasert, W.

    2014-01-01

    Establishing in-vitro dose calibration curves is important for reconstruction of radiation dose in the exposed individuals. The aim of this pioneering work in Thailand was to generate dose-response curves using conventional biological dosimetry: dicentric chromosome assay (DCA) and premature chromosome condensation (PCC) assay. The peripheral blood lymphocytes were irradiated with 137 Cs at a dose rate of 0.652 Gy/min to doses of 0.1, 0.25, 0.5, 0.75, 1, 2, 3, 4 and 5 Gy for DCA technique, and 5, 10, 15, 20 and 25 Gy for PCC technique. The blood samples were cultured and processed following the standard procedure given by the IAEA with slight modifications. At least 500-1,000 metaphases or 100 dicentrics/ PCC rings were analyzed using an automated metaphase finder system. The yield of dicentrics with dose was fitted to a linear quadratic model using Chromosome Aberration Calculation Software (CABAS, version 2.0), whereas the dose-response curve of PCC rings was fitted to a linear relationship. These curves will be useful for in-vitro dose reconstruction and can support the preparedness for radiological emergency in the country.

  18. Analysis of complex-type chromosome exchanges in astronauts' lymphocytes after space flight as a biomarker of high-LET exposure

    Science.gov (United States)

    George, Kerry; Wu, Honglu; Willingham, Veronica; Cucinotta, Francis A.

    2002-01-01

    High-LET radiation is more efficient in producing complex-type chromosome exchanges than sparsely ionizing radiation, and this can potentially be used as a biomarker of radiation quality. To investigate if complex chromosome exchanges are induced by the high-LET component of space radiation exposure, damage was assessed in astronauts' blood lymphocytes before and after long duration missions of 3-4 months. The frequency of simple translocations increased significantly for most of the crewmembers studied. However, there were few complex exchanges detected and only one crewmember had a significant increase after flight. It has been suggested that the yield of complex chromosome damage could be underestimated when analyzing metaphase cells collected at one time point after irradiation, and analysis of chemically-induced PCC may be more accurate since problems with complicated cell-cycle delays are avoided. However, in this case the yields of chromosome damage were similar for metaphase and PCC analysis of astronauts' lymphocytes. It appears that the use of complex-type exchanges as biomarker of radiation quality in vivo after low-dose chronic exposure in mixed radiation fields is hampered by statistical uncertainties.

  19. Persistence of chromatid damage after G2 phase X-irradiation in lymphoblastoid cells from Gardner's syndrome

    International Nuclear Information System (INIS)

    Takai, Setsuo; Price, F.M.; Sanford, K.K.; Tarone, R.E.; Parshad, Ram

    1990-01-01

    Previous reports showed that skin fibroblasts or peripheral blood lymphocytes from individuals with hereditary cancer or with a genetic disorder predisposing to cancer show an abnormally high frequency of chromatid damage after X-irradiation in G 2 phase. The reproducibility of this response suggested that it could provide the basis of an assay for genetic predisposition to cancer. The present blind study tested whether lymphoblastoid cell lines could also be used in this assay. Lymphoblastoid cell lines from patients with Gardner's syndrome (GS) were compared with those from clinically normal controls. In metaphase cells collected during the first 30 min after X-irradiation (58R), frequencies of chromatid breaks and gaps were similar in GS and normal cells. However, in metaphase cells collected from 0.5 to 1.5 h and 1.5 to 2.5 h after X-irradiation, the total unrepaired damage for each GS cell line was greater than that observed in any of the lines from clinically normal controls. The persistence of chromatid damage in the GS cells after X-irradiation suggests a deficiency or imbalance in the repair or processing of the radiation-induced DNA damage. The results show that lymphoblastoid cell lines in early passage can be used in this cytogenetic assay to identify members in a GS family who have the GS gene(s) or other individuals with a genetic predisposition to cancer. (author)

  20. Karyotype, heterochromatin distribution and meiosis of Zabrotes subfasciatus (Bohemann) (Coleoptera: Chrysomelidae, Bruchinae)

    International Nuclear Information System (INIS)

    Correa, Ronan X.; Santos, Igor S.; Silva, Janisete G.; Costa, Marco A.; Pompolo, Silvia G.

    2008-01-01

    Zabrotes subfasciatus (Boh.) has been extensively studied in its agronomic and biochemical aspects due to its importance as a damaging insect to leguminous grains during storage. The few cytogenetic studies published on this species yielded conflicting results. In this study, the karyotype was analyzed in order to accurately describe the chromosome C-banding patterns and meiosis. The brain ganglion at the pre pupa and the adult and pupal testes were analyzed. All individuals had 26 chromosomes in both brain ganglion and spermatogonic mitotic metaphases. These chromosomes were classified as follows: the 12 th pair and the Y chromosome were telocentric; the X chromosome was acrocentric; the 4 th and 5 th pairs were sub metacentric; and the remaining pairs were all metacentric. One of the members of the 5 th pair presented a secondary constriction. All chromosomes presented pericentromeric heterochromatin. The large arms of the pairs 5, 9 and X presented heterochromatin. The X chromosome showed to be heteropyknotic throughout the prophase of the fi rst meiotic division. The sub phases of prophase I were atypical and meiosis II was rarely identified. Testes of all males showed a few cells; the bivalents were rod-like shaped in metaphase I. Karyological formulae were 2n = 24 + XX in females and 2n = 24 + XYp and either n = 12 + X or n = 12 + Y in males. (author)

  1. Karyotype, heterochromatin distribution and meiosis of Zabrotes subfasciatus (Bohemann) (Coleoptera: Chrysomelidae, Bruchinae)

    Energy Technology Data Exchange (ETDEWEB)

    Correa, Ronan X.; Santos, Igor S.; Silva, Janisete G.; Costa, Marco A. [Universidade Estadual de Santa Cruz, Ilheus, BA (Brazil). Dept. de Ciencias Biologicas; Pompolo, Silvia G. [Universidade Federal de Vicosa, MG (Brazil). Dept. de Biologia Geral

    2008-09-15

    Zabrotes subfasciatus (Boh.) has been extensively studied in its agronomic and biochemical aspects due to its importance as a damaging insect to leguminous grains during storage. The few cytogenetic studies published on this species yielded conflicting results. In this study, the karyotype was analyzed in order to accurately describe the chromosome C-banding patterns and meiosis. The brain ganglion at the pre pupa and the adult and pupal testes were analyzed. All individuals had 26 chromosomes in both brain ganglion and spermatogonic mitotic metaphases. These chromosomes were classified as follows: the 12{sup th} pair and the Y chromosome were telocentric; the X chromosome was acrocentric; the 4{sup th} and 5{sup th} pairs were sub metacentric; and the remaining pairs were all metacentric. One of the members of the 5{sup th} pair presented a secondary constriction. All chromosomes presented pericentromeric heterochromatin. The large arms of the pairs 5, 9 and X presented heterochromatin. The X chromosome showed to be heteropyknotic throughout the prophase of the fi rst meiotic division. The sub phases of prophase I were atypical and meiosis II was rarely identified. Testes of all males showed a few cells; the bivalents were rod-like shaped in metaphase I. Karyological formulae were 2n = 24 + XX in females and 2n = 24 + XYp and either n = 12 + X or n = 12 + Y in males. (author)

  2. In vitro growth and maturation of isolated caprine preantral follicles: Influence of insulin and FSH concentration, culture dish, coculture, and oocyte size on meiotic resumption.

    Science.gov (United States)

    Silva, G M; Brito, I R; Sales, A D; Aguiar, F L N; Duarte, A B G; Araújo, V R; Vieira, L A; Magalhães-Padilha, D M; Lima, L F; Alves, B G; Silveira, L B R; Lo Turco, E G; Rodrigues, A P; Campello, C C; Wheeler, M B; Figueiredo, J R

    2017-03-01

    The aims of this study were: (1) to evaluate the effect of different insulin concentrations, alone or in combination with either a fixed FSH concentration or increasing FSH concentrations on the in vitro culture of isolated caprine preantral follicles and (2) to analyze the efficiency of two IVM media and maturation culture systems (with or without coculture with in vivo grown oocytes) on the meiosis resumption. Secondary follicles were cultured for 18 days in a basic medium supplemented with low- or high-insulin concentration alone or with a fixed FSH concentration or with increasing FSH concentrations. Oocytes grown in vivo or in vitro were matured alone or cocultured. The high-insulin concentration associated with fixed FSH treatment had higher meiotic resumption rate (P rates of germinal vesicle, germinal vesicle breakdown, metaphase I, metaphase II (MII), meiotic resumption, and oocyte diameter were similar between the maturation media. In conclusion, a basic medium supplemented with 10-μg/mL insulin and 100-μg/mL FSH throughout the culture period improved meiotic resumption rate and produced MII oocytes from caprine preantral follicles cultured in vitro. The MII rate was similar between in vivo and in vitro grown oocytes ≥110 μm. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. MAPK-activated protein kinase 2 is required for mouse meiotic spindle assembly and kinetochore-microtubule attachment.

    Directory of Open Access Journals (Sweden)

    Ju Yuan

    Full Text Available MAPK-activated protein kinase 2 (MK2, a direct substrate of p38 MAPK, plays key roles in multiple physiological functions in mitosis. Here, we show for the first time the unique distribution pattern of MK2 in meiosis. Phospho-MK2 was localized on bipolar spindle minus ends and along the interstitial axes of homologous chromosomes extending over centromere regions and arm regions at metaphase of first meiosis (MI stage in mouse oocytes. At metaphase of second meiosis (MII stage, p-MK2 was localized on the bipolar spindle minus ends and at the inner centromere region of sister chromatids as dots. Knockdown or inhibition of MK2 resulted in spindle defects. Spindles were surrounded by irregular nondisjunction chromosomes, which were arranged in an amphitelic or syntelic/monotelic manner, or chromosomes detached from the spindles. Kinetochore-microtubule attachments were impaired in MK2-deficient oocytes because spindle microtubules became unstable in response to cold treatment. In addition, homologous chromosome segregation and meiosis progression were inhibited in these oocytes. Our data suggest that MK2 may be essential for functional meiotic bipolar spindle formation, chromosome segregation and proper kinetochore-microtubule attachments.

  4. Optimasi Penambahan Colcemid pada Karyotyping Kultur Mecenchymal Stem Cells (MSC Mencit

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    Ratih Rinendyaputri

    2016-02-01

    Full Text Available AbstractControl of the genetic stability of stem cells prior to the conduct of therapy is essential to prevent effects such as stem cell transformation. Karyotyping is a conventional technique to conduct an analysis of the number and structure of chromosomes. The analysis can only be performed on metaphase stage that needs to be optimized to get the cell at that stage because the length of the cell cycle are different in the each cell types. This study aims to obtain an optimal time to get MSC at metaphase stage. The study was conducted at the stem cell laboratory of Center for Biomedical and Basic Technology of Health. The event begins with isolation using flushing technique at the femur and tibia of mice. Furthermore, the culture in vitro and induction colcemid 0,25μg/ml for 8,16 and 24 hours to get the MSC at metaphase stage. KCl solution with a concentration of 0.075 M and 0,045 M used as a solvent hipotonis. Results showed that 16 hours of induction colcemid 0,25μg/ml in 0.075 M KCl solution usage percentage of MSC who are at metaphase stage and do the highest analysis (p<0.05. In this study 16 hours induction colcemid 0,25μg/ml is the optimal time to obtain metaphase stage of the MSC from bone marrow of mice.Keywords: mecenchymal stem cell, karyotyping, colcemidAbstrakKontrol terhadap stabilitas genetik pada sel punca sebelum pelaksanan terapi merupakan hal yang penting untuk mencegah efek seperti transformasi sel punca yang dapat terjadi. Secara konvensional dapat dilakukan karyotyping untuk melakukan analisis terhadap jumlah dan struktur kromosom. Analisis hanya dapat dilakukan pada tahap metafase sehingga perlu dilakukan optimasi untuk mendapatkan sel pada tahap tersebut mengingat panjang siklus sel setiap jenis sel berbeda. Penelitian ini bertujuan untuk memperoleh waktu yang optimal untuk mendapatkan MSC pada tahap metafase. Penelitian dilakukan di Laboratorium stem cell Pusat Biomedis dan Teknologi Dasar Kesehatan Badan Litbangkes

  5. Análise citológica e cariométrica da ação da colchina sôbre a espermatogênese dos hemípteros Cytologic and caryometric analysis of the action of colchicine on the spermatogenesis in hemiptera

    Directory of Open Access Journals (Sweden)

    G. Schreiber

    1951-03-01

    Full Text Available The action of colchicine upon the spermatogenesis of Triatoma infestans, (Hemipt. Heteroptera, has been studied and the different categories of giant spermatids that appear during the treatment have been compared with the nuclear volumes of the whole series of normal spermatogenetic stages. The following facts have been ascertained: 1 4 hours after the treatment the gonial mitotic metaphases, and the 1st. and 2nd. metaphases of meiosis are stopped. The prophasic stages of meiosis and diakynesis appear to be normal. After 9 days of treatment, all the tetrads are broken in the meiotic metaphases and the cells appear with 44 and 22 chromosomes respectively, scattered in the cytoplasm. 2 At 9 days, practically all spermatogenetic stages have disappeared except for a few cysts of spermatogonia, and practically the whole testicle is full of cysts of spermatozoa and spermatid, with some large zones of necrosis with pycnotic nuclei. The spermatids appear to be of different sizes and the statistical analysis of the nuclear volumes gives a polymodal hystogram with 4 modes, whose volumes are in the ratio of 1:2:4:8. Ripe spermatozoa seem to have a certain volume variability, that has not been possible to analyse quantitatively. All these facts confirm what DOOLEY found in the colchicinized Orthoptera testicle. 3 The caryometric analysis conducted statistically on the normal stages of the spermatogenesis (resting spermatogonia, gonial prophases, leptotene, "confused stage", diakynesis, and spermatid revealed the following facts: a Considering the volume of the resting, spermatogonia as 1, their mitotic prophases have a volume of 2. Some rare prophases appear to have a volume of 4 and probably belong to tetraployd spermatogonia normally present in the testicle of Hemiptera. b The first spermatocyte at the beginning of the auxocitary growth (leptotene has a volume of 2, which is equal to that of them gonial prophase. It grows further during the "confused

  6. A descriptive mutagenicity assessment of tretinoin in Allium sativum

    International Nuclear Information System (INIS)

    Dela Llana, Jonamine M.; Reyes, Florence C.

    1999-01-01

    This paper is primarily designed to assess the mutagenicity of tretinoin by applying the Allium test. Furthermore, it has the following objectives: to evaluate the macroscopic abnormalities caused by tretinoin based on root length and root form parameters; to investigate whether tretinoin can induce aberrances in cell division such as the formation of micronucleus, anaphase bridges, early anaphase, C-metaphase, sticky chromosome, stretched chromosome, vagrant chromosome and precocious chromosome; to determine the variation in the aberrations in the different concentration of tretinoin. Procedure: eight hundred equal-sized garlic bulbs were immersed in various concentrations of tretinoin and in tap water as control. These were divided into two groups. Six hundred bulbs were evaluated for macroscopic parameters while the remaining two hundred bulbs were fixed for microscopic observations. The Allium test set-ups were placed in the plant laboratory of UP-Manila. The were harvested on the third and on the fifth day. The fixed roots were examined in the Cytogenetics Laboratory of the Philippine Nuclear Research Institute. The data gathered for macroscopic parameter was statistically tested using Complete Randomized Design and the Tukey's Honestly Significant Difference. The microscopic abnormalities were determined descriptively for every concentration. Findings: analysis of macroscopic and microscopic parameters showed that: according to the analyses of variances, the number of roots, the root length and the number of root forms such as straight, bent, bulbous and tapered were not equal in all concentrations. However, the difference in the number of curled roots was not significant.; the root length distinctly showed the toxicity effect of tretinoin. The growth or the length of roots decreases as the tretinoin concentration increases; the mitotic abnormalities observed in the garlic cells include micronucleus, anaphase bridges, early anaphase. C-metaphase, sticky

  7. Establishment of conditions for ovum pick up and IVM of jennies oocytes toward the setting up of efficient IVF and in vitro embryos culture procedures in donkey (Equus asinus).

    Science.gov (United States)

    Goudet, Ghylène; Douet, Cécile; Kaabouba-Escurier, Aurore; Couty, Isabelle; Moros-Nicolás, Carla; Barrière, Philippe; Blard, Thierry; Reigner, Fabrice; Deleuze, Stefan; Magistrini, Michèle

    2016-07-15

    Most wild and domestic donkey breeds are currently endangered or threatened. Their preservation includes the creation of a Genome Resource Bank. Embryos cryopreservation allows the preservation of genetics from both male and female and is the fastest method to restore a breed. Because embryo production in vivo is limited in equids, our objective was to establish conditions for in vitro production of embryos in donkey using ovum pick up (OPU), IVM, IVF, and in vitro culture of zygotes. Donkey cumulus-oocyte complexes (COCs) were collected by transvaginal ultrasound-guided aspirations OPU in adult cyclic jennies and in vitro matured in tissue culture medium 199 supplemented with fetal calf serum and epidermal growth factor for 24, 30, 34, or 38 hours. They were preincubated with oviductal fluid for 30 minutes, coincubated with frozen-thawed donkey semen treated with procaine for 18 hours, and cultured for 30 hours in Dulbecco's Modified Eagle Medium-F12 supplemented with NaHCO3, fetal calf serum, and gentamycin. From the five OPU sessions, we collected 92 COCs in 193 follicles (48%) with an average of 4.2 COCs per jenny. All COCs were expanded after more than 24-hour IVM. At collection, jennies oocytes contained a germinal vesicle. Metaphase 1 oocytes were observed after 30-hour IVM and 44% were in metaphase 2 after 34-hour IVM. In our conditions, IVM of donkey oocytes was slower than IVM of equine oocytes and optimal duration for donkey oocytes IVM may be 34 hours. Only 15% of jennies oocytes contained two pronuclei after coincubation with donkey spermatozoa and none of them developed further after 48 hours post-IVF. Moreover, some parthenogenetic activation occurred. Thus, the treatment of donkey sperm with procaine may not be efficient for IVF. In conclusion, we established for the first time conditions for OPU in jennies with high recovery rates. We reported that IVM of jennies oocytes can produce 44% of metaphase 2 oocytes after 34 hours in culture

  8. Upregulation of meiosis-specific genes in lymphoma cell lines following genotoxic insult and induction of mitotic catastrophe

    Directory of Open Access Journals (Sweden)

    Cragg Mark S

    2006-01-01

    Full Text Available Abstract Background We have previously reported that p53 mutated radioresistant lymphoma cell lines undergo mitotic catastrophe after irradiation, resulting in metaphase arrest and the generation of endopolyploid cells. A proportion of these endopolyploid cells then undergo a process of de-polyploidisation, stages of which are partially reminiscent of meiotic prophase. Furthermore, expression of meiosis-specific proteins of the cancer/testis antigens group of genes has previously been reported in tumours. We therefore investigated whether expression of meiosis-specific genes was associated with the polyploidy response in our tumour model. Methods Three lymphoma cell lines, Namalwa, WI-L2-NS and TK6, of varying p53 status were exposed to a single 10 Gy dose of gamma radiation and their responses assessed over an extended time course. DNA flow cytometry and mitotic counts were used to assess the kinetics and extent of polyploidisation and mitotic progression. Expression of meiotic genes was analysed using RT-PCR and western blotting. In addition, localisation of the meiotic cohesin REC8 and its relation to centromeres was analysed by immunofluorescence. Results The principal meiotic regulator MOS was found to be significantly post-transcriptionally up-regulated after irradiation in p53 mutated but not p53 wild-type lymphoma cells. The maximum expression of MOS coincided with the maximal fraction of metaphase arrested cells and was directly proportional to both the extent of the arrest and the number of endopolyploid cells that subsequently emerged. The meiotic cohesin REC8 was also found to be up-regulated after irradiation, linking sister chromatid centromeres in the metaphase-arrested and subsequent giant cells. Finally, RT-PCR revealed expression of the meiosis-prophase genes, DMC1, STAG3, SYCP3 and SYCP1. Conclusion We conclude that multiple meiotic genes are aberrantly activated during mitotic catastrophe in p53 mutated lymphoma cells after

  9. Analysis of the karyotype of expanded human adipose-derived stem cells for bone reconstruction of the maxillo-facial region.

    Science.gov (United States)

    Bellotti, C; Stanco, D; Ragazzini, S; Romagnoli, L; Martella, E; Lazzati, S; Marchetti, C; Donati, D; Lucarelli, E

    2013-01-01

    Mesenchymal stem cells (MSC) and adipose-derived stem cells (ASC) were recently proposed for bone maxillofacial reconstruction in association with biomaterials. For this application MSC must be ex-vivo expanded in order to obtain, for a given volume of implanted biomaterial, a relevant number of bone forming cells. Previously conducted pre-clinical studies suggested that a concentration of 6 x 10(8) ASC associated with 900 mg of anorganic bovine bone (ABB) could be effective for human maxillary sinus floor elevation. A keystone issue to guarantee the quality and safety of Advanced Therapy Medicinal Products containing expanded MSC and ASC is their chromosome stability in culture: this topic has been widely investigated and conflicting results have been published. Abnormal karyotype of human ex-vivo expanded MSC and ASC was found by some authors, while, at the same time, several other studies showed the MSC and ASC karyotype to be normal. It is therefore important that all the results obtained on MSC and ASC karyotype analysis be published. Given this context, the aim of this manuscript, aim of this manuscript is to verify the karyotype stability of ASC in view of their applications in clinical trials. ASC obtained from the adipose tissue of 4 donors were expanded over extended culture time. Based on previous ASC expansions we hypothesized to be able to obtain 6 x 10(8) cells by passage 7. Karyotype analysis of 30 metaphases was planned to be investigated at passage 2, 7, and 15 in all the cultures. No abnormalities were found in the karyotype of two donors at all the passages tested, while a translocation was found in 2 metaphases of a donor at passage 7, but not at passage 15, and in the fourth donor in 5 metaphases a trisomy was found at passage 15. Chromosomal abnormalities were detected only after extended ASC expansion. Whether these anomalies can be related to risk for the patient's safety will have to be demonstrated by in-vivo studies.

  10. [The detection of living triploid of Haliotis discus hannai--directly making chromosome sample of mantle and epipedium tentacle].

    Science.gov (United States)

    Li, Ya-Juan; Mao, Lian-Ju; Li, Xia; Wang, Zi-Chen; Song, Jian

    2002-06-01

    Ploidy detection method of living body is a very important component part in polyploid breeding. There were few reports about chromosome ploidy detection of adult abalone (Haliotis discus hannai), except preparing adult abalone chromosome by means of chopping method using gill tissues and squashing method using gonad, also with these methods the abalone should be killed for getting these tissues. In polyploid breeding living triploid abalones are needed, so ploidy detection technique of living abalone is especially important. In this paper the orthogonal experiment of three factors and three levels L9(3(4)) using mantle and epipodium tentacle as material of living tissues which were vido immersing in PHA solution for preparing chromosome and count the frequence of cell division was first reported. Three factors and three levels were raising temperature: 16 degrees C, 18 degrees C, 20 degrees C; PHA concentration: 0.5%, 1.0%, 1.5%; sampling time: 5:00-6:00, 17:00-18:00, 23:00-24:00, respectively. The tests were repeated two times. The test results showed that the optimal patterns effecting the mitotic frequency of three factors and three levels in triploid abalone detection of chromosome from mantles and epipodium of living bodies were: raising temperature 20 degrees C, PHA concentration 1.0%, and sampling time 17:00-18:00. The order of three factors was: raising temperature-->PHA concentration-->sampling time. Under optimal pattern of three factors and three levels many clear, proper in length and well-scattered metaphase chromosomes could be obtained. The advantages of preparing chromosome using mantles and epipodium tentacle were sampling convenient, no body size restriction and no influence of survival of abalone. After using PHA treated tissues, mitotic frequency could be improved and 1.00% PHA concentration was the best. The key of chromosome preparation was the cells in mitotic metaphase, so the time of sampling was important. During 17:00-18:00, sampling of

  11. Chromosome dosimetry: the influence of culture media on the proliferation of irradiated and unirradiated human lymphocytes

    International Nuclear Information System (INIS)

    Purrott, R.J.; Lloyd, D.C.; Vulpis, N.

    1981-01-01

    The proliferation of phytohaemagglutinin stimulated human lymphocytes in four types of synthetic culture medium has been studied using the fluorescence plus Giemsa staining technique to determine cell cycle status. 48 hour cultures of unirradiated cells containing Ham's F10 or RPMI 1640 media yielded significant numbers of second cycle metaphases. Cultures containing Eagle's MEM or TC 199 media, however, required longer incubation times to produce appreciable numbers of second division cells. Intrinsic differences between donors in the rate of proliferation had little effect on the relative ranking of the media. Radiation induced mitotic delay of about 1 hour per Gray was observed for each medium. The relevance of these results to the accuracy of radiation dose estimation by chromosome aberration analysis is discussed. (author)

  12. Chromosome aberration analysis in peripheral lymphocytes of Gulf war and Balkans war veterans

    International Nuclear Information System (INIS)

    Schroeder, H.; Heimers, A.; Frentzel-Beyme, R.; Schott, A.; Hoffmann, W.

    2003-01-01

    Chromosome aberrations and sister chromatid exchanges (SCEs) were determined in standard peripheral lymphocyte metaphase preparations of 13 British Gulf War veterans, two veterans of the recent war in the Balkans, and one veteran of both wars. All 16 volunteers suspect exposures to depleted uranium while deployed at the two different theatres of war in 1990 and later on. The Bremen laboratory control served as a reference in this study. Compared with this control there was a statistically significant increase in the frequency of dicentric chromosomes (dic) and centric ring chromosomes (cR) in the veterans' group, indicating a previous exposure to ionising radiation. The statistically significant overdispersion of dic and cR indicates non-uniform irradiation as would be expected after non-uniform exposure and/or exposure to radiation with a high linear energy transfer. The frequency of SCEs was decreased when compared with the laboratory control. (author)

  13. Discrimination of chromosome by autoradiography

    International Nuclear Information System (INIS)

    Masubuchi, Masanori

    1975-01-01

    This paper describes discrimination of chromosome by autoradiography. In this method, the difference in DNA synthetic phase between each chromosome was used as a standard, and the used chromosome was in metaphase, as morphological characteristics were markedly in this phase. Cell cycle and autoradiography with 3 H-thymidine were also examined. In order to discriminate chromosome by autoradiography, it was effective to utilize the labelled pattern in late DNA synthetic phase, where asynchronous replication of chromosome appeared most obviously. DNA synthesis in chromosome was examined in each DNA synthetic phase by culturing the chromosome after the treatment with 3 H-thymidine and altering the time to prepare chromosome specimen. Discrimination of chromosome in plants and animals by autoradiography was also mentioned. It was noticed as a structural and functional discrimination of chromosome to observe amino acid uptake into chromosome protein and to utilize the difference in labelled pattern between the sites of chromosome. (K. Serizawa)

  14. A Cdk1 phosphomimic mutant of MCAK impairs microtubule end recognition

    Directory of Open Access Journals (Sweden)

    Hannah R. Belsham

    2017-12-01

    Full Text Available The microtubule depolymerising kinesin-13, MCAK, is phosphorylated at residue T537 by Cdk1. This is the only known phosphorylation site within MCAK’s motor domain. To understand the impact of phosphorylation by Cdk1 on microtubule depolymerisation activity, we have investigated the molecular mechanism of the phosphomimic mutant T537E. This mutant significantly impairs microtubule depolymerisation activity and when transfected into cells causes metaphase arrest and misaligned chromosomes. We show that the molecular mechanism underlying the reduced depolymerisation activity of this phosphomimic mutant is an inability to recognise the microtubule end. The microtubule-end residence time is reduced relative to wild-type MCAK, whereas the lattice residence time is unchanged by the phosphomimic mutation. Further, the microtubule-end specific stimulation of ADP dissociation, characteristic of MCAK, is abolished by this mutation. Our data shows that T537E is unable to distinguish between the microtubule end and the microtubule lattice.

  15. EFFECT OF CYSTEAMINE ON THE RATE OF IN VITRO MATURATION OF OOCYTES IN TWO MEDIA

    Directory of Open Access Journals (Sweden)

    A. Mohammadi-Rousheh

    2006-07-01

    Full Text Available Rate of in vitro maturation of oocytes is one of the challenges of assisted reproductive techniques. In this study we investigated the effects of supplementation of cysteamine on the rate of in vitro maturation of oocytes in two different media. Germinal vesicle oocytes were collected from mouse ovary and cultured in two media (TCM199 and MEME with 0, 50, 100, 200, 500 µM/ml cysteamine. Number of germinal vesicle breakdowns and metaphase II oocytes were recorded. The results showed that the rate of in vitro maturation in 100 µM/ml cysteamine was significantly higher compared to control (P < 0.05. Evaluation of two media in this study showed that TCM199 improved the rate of in vitro maturation and oocyte maturation better than MEME; however, this difference was not statistically significant. These findings indicate that TCM199 as compared to MEME was better in rate of in vitro maturation of oocytes.

  16. Nonrandom distribuion of chromosome breaks in cultured lymphocytes of normal subjects

    Energy Technology Data Exchange (ETDEWEB)

    Ayme, S.; Mattei, J.F.; Mattei, M.G.; Aurran, Y.; Giraud, F.

    1976-02-29

    Breakpoint distribution was studied from cultured lymphocytes on 7653 metaphases from 524 subjects whose karyotypes were normal. The mean break rate was 5% in both sexes. The frequency increased significantly after 40 years and varied during the year. The location of the breaks was very different from the expected random distribution. The break frequency for each chromosome was different according to the type of break (chromatid, simple chromosomal and chromosomal involving rearrangements). The location of the breaks was also studied according to the type of band and with respect to the centromere. A comparison between spontaneous breaks, x-ray induced breaks, breaks in Fanconi's anemia and in congenital rearrangements, show very significant differences.

  17. Multi-micronucleus cells related with viral diseases, detected in the study of children affected by the Chernobyl accident

    International Nuclear Information System (INIS)

    Garcia L, O.; Lamadrid, A.I.; Manzano, J.

    1996-01-01

    Cells with multiple chromosome aberrations have been observed in human peripheral blood lymphocytes. Different explanation have proposed, included hot particle induction in persons related to the Chernobyl accident. The frequency of chromosome aberration and micronuclei were established in 14 Ukrainian children with different hematological disorders. They arrived in Cuba thanks to the program by means of which medical attention is offered to children from areas affected by the Chernobyl accident. At least 500 metaphases and bi-nucleate cells were analyzed in each case. The detection of 4 cells with 7-11 micronuclei in a 14 year old boy with cat scratch disease was the most significant cytogenetical finding. The viral origin of the cat scratch disease has been reported, this suggested a viral etiology of the cells with multiple micronuclei. No rogue cells were detected. Cells with multiple micronuclei or rogue cells were not found in other patients from this group. (authors). 7 refs., 3 tabs

  18. Chromosome numbers, meiotic behavior and pollen fertility in a collection of Paspalum nicorae Parodi accessions

    Directory of Open Access Journals (Sweden)

    Camila Aparecida de Oliveira dos Reis

    2008-01-01

    Full Text Available Chromosome number, meiotic behavior and pollen viability were evaluated in a collection of 53 Paspalumnicorae Parodi accessions, which are part of a breeding project of the species. All accessions are tetraploid, with 2n=4x=40.Despite the invariable chromosome numbers, there was variation among accessions in the frequencies of different chromosomeconfigurations at diakinesis and metaphase I, such as univalents, trivalents and quadrivalents. Other abnormalities asbridges and laggards were also observed at anaphase and telophase I. Meiotic indexes ranged from 82.00 to 99.50% andpollen viability from 88.99 to 95.06%. As the species is pseudogamous apomictic, fertile pollen is necessary for endospermformation. Results show that all plants are meiotically stable and have enough fertile pollen to be used as male parents incontrolled crosses.

  19. Centromeric heterochromatin: the primordial segregation machine.

    Science.gov (United States)

    Bloom, Kerry S

    2014-01-01

    Centromeres are specialized domains of heterochromatin that provide the foundation for the kinetochore. Centromeric heterochromatin is characterized by specific histone modifications, a centromere-specific histone H3 variant (CENP-A), and the enrichment of cohesin, condensin, and topoisomerase II. Centromere DNA varies orders of magnitude in size from 125 bp (budding yeast) to several megabases (human). In metaphase, sister kinetochores on the surface of replicated chromosomes face away from each other, where they establish microtubule attachment and bi-orientation. Despite the disparity in centromere size, the distance between separated sister kinetochores is remarkably conserved (approximately 1 μm) throughout phylogeny. The centromere functions as a molecular spring that resists microtubule-based extensional forces in mitosis. This review explores the physical properties of DNA in order to understand how the molecular spring is built and how it contributes to the fidelity of chromosome segregation.

  20. Interactive Karyotyping Training

    Directory of Open Access Journals (Sweden)

    Ashwin Kotwaliwale

    2013-01-01

    Full Text Available Despite the wide use of newer techniques in genetic diagnostics, there remains a need for technologists to learn human chromosome morphology, identify abnormal metaphases and report clinical abnormalities. Global short age of cytogenetic trainers and a time consuming training process makes Karyotyping training difficult. We have developed a web based interactive Karyotyping training tool, KaryoTutor©, that allows technologists to learn karyotyping in an interactive environment and aids the trainer in the training process. KaryoTutor©provides visual clues for identifying abnormal chromosomes, provides instant test scores and includes a reference library of ideograms,sample chromosome images and reference materials. Trainees are able to recursively work on a case till a satisfactory result is achieved,with KaryoTutor providing interactive inputs.Additionally, trainers can assign cases and monitor trainee progress using audit trail management and other administrative features.

  1. Time-sequential observation of spindle and phragmoplast orientation in BY-2 cells with altered cortical actin microfilament patterning.

    Science.gov (United States)

    Kojo, Kei H; Yasuhara, Hiroki; Hasezawa, Seiichiro

    2014-06-18

    Precise division plane determination is essential for plant development. At metaphase, a dense actin microfilament meshwork appears on both sides of the cell center, forming a characteristic cortical actin microfilament twin peak pattern in BY-2 cells. We previously reported a strong correlation between altered cortical actin microfilament patterning and an oblique mitotic spindle orientation, implying that these actin microfilament twin peaks play a role in the regulation of mitotic spindle orientation. In the present study, time-sequential observation was used to reveal the progression from oblique phragmoplast to oblique cell plate orientation in cells with altered cortical actin microfilament patterning. In contrast to cells with normal actin microfilament twin peaks, oblique phragmoplast reorientation was rarely observed in cells with altered cortical actin microfilament patterning. These results support the important roles of cortical actin microfilament patterning in division plane orientation.

  2. A remarkable autosomal heteromorphism in Pseudoryzomys simplex 2n = 56; FN = 54-55 (Rodentia, Sigmodontinae

    Directory of Open Access Journals (Sweden)

    Camila Nascimento Moreira

    2013-01-01

    Full Text Available Pseudoryzomys simplex, the false rice rat, is a monotypic genus of the Oryzomyini tribe (Sigmodontinae distributed in part of Bolivia, Paraguay, Argentina and Brazil. Its diploid number has been described as 56 acrocentric chromosomes decreasing in size and no karyotype figure has been depicted. Herein, we present karyotypic data on P. simplex, including chromosome banding and molecular fluorescent in situ hybridization using telomeric sequences and the whole X-chromosome of its sister clade Holochilus brasiliensis (HBR as probes. A case of remarkable autosomal heteromorphism due to the presence of a whole heterochromatic arm leading to the variability of FN is reported, as well as the occurrence of regions of homology between the X and Y chromosomes (pseudoautosomal regions after chromosome painting with the HBR X probe on P. simplex metaphases.

  3. Abnormal spindle orientation during microsporogenesis in an interspecific Brachiaria (Gramineae hybrid

    Directory of Open Access Journals (Sweden)

    Andréa Beatriz Mendes-Bonato

    2006-01-01

    Full Text Available This paper reports a case of abnormal spindle orientation during microsporogenesis in an interspecific hybrid of the tropical grass Brachiaria. In the affected plant, prophase I was normal. In metaphase I, bivalents were regularly co-oriented but distantly positioned and spread over the equatorial plate. In anaphase I, chromosomes failed to converge into focused poles due to parallel spindle fibers. As a consequence, in telophase I, an elongated nucleus or several micronuclei were observed in each pole. In the second division, the behavior was the same, leading to polyads with several micronuclei. A total of 40% of meiotic products were affected. The use of this hybrid in production systems needing good-quality seeds is discussed.

  4. Avaliação da maturação nuclear in vitro de oócitos de gatas domésticas (Felis catus pré-púberes e púberes

    Directory of Open Access Journals (Sweden)

    Maria Denise Lopes

    2004-08-01

    Full Text Available This study assessed the in vitro oocyte nuclear maturation in adult and young domestic cats. Fifteen ovaries were used; 10 from adult females and 5 from young females. The oocytes collected by aspiration were quantified and classified. The oocytes classified as excellent and/or regular were grouped (10 oocyte /drop in culture medium covered with mineral oil in disposable Petri dishes. The oocyte were incubated at 38°C and 5% de CO2 for 48 hours and then washed with 0,4% hyaluronidase, fixed in methanol/acetic acid and stained with acetic orcein. Evaluation of chromosomal configuration of oocytes matured in vitro showed 44,68% of oocyte in metaphase II in the adult female group and 25,32% in the young donor group, showing that puberty influences oocyte capacity for in vitro development.

  5. Cytogenetic assessment of the effect of radioecological situation to the residents of Zhovti Vody city

    Directory of Open Access Journals (Sweden)

    L. K. Bezdrobna

    2016-12-01

    Full Text Available Results of chromosomal aberration frequency and the spectrum study in peripheral blood lymphocytes cultures of the residents at age of 19 - 60 in Zhovti Vody city (n = 42 and Kyiv (n = 42, who did not have professional contact with radiation and others mutagenic factors, and also the incidence analysis of adult population of these cities and Ukraine in total are given. Average frequency of metaphases with chromosomal aberration and chromosome type aberrations specific to the action of the radiation factor, namely unstable interchromosomal exchanges with the accompanying fragment and without fragment, free acentrics, atypical monocentrics and chromatid type aberrations, that represent the total genome instability, is significantly higher in the group of Zhovti Vody's residents comparing to the group of Kyiv’s residents. Futhermore, multiaberrant cells in three persons from Zhovti Vody were found. The morbidity of malignant tumors in Zhovti Vody is higher than in Kyiv and in Ukraine in total.

  6. Mitotic spindle proteomics in Chinese hamster ovary cells.

    Directory of Open Access Journals (Sweden)

    Mary Kate Bonner

    Full Text Available Mitosis is a fundamental process in the development of all organisms. The mitotic spindle guides the cell through mitosis as it mediates the segregation of chromosomes, the orientation of the cleavage furrow, and the progression of cell division. Birth defects and tissue-specific cancers often result from abnormalities in mitotic events. Here, we report a proteomic study of the mitotic spindle from Chinese Hamster Ovary (CHO cells. Four different isolations of metaphase spindles were subjected to Multi-dimensional Protein Identification Technology (MudPIT analysis and tandem mass spectrometry. We identified 1155 proteins and used Gene Ontology (GO analysis to categorize proteins into cellular component groups. We then compared our data to the previously published CHO midbody proteome and identified proteins that are unique to the CHO spindle. Our data represent the first mitotic spindle proteome in CHO cells, which augments the list of mitotic spindle components from mammalian cells.

  7. Microfluidic bioreactors for culture of non-adherent cells

    DEFF Research Database (Denmark)

    Shah, Pranjul Jaykumar; Vedarethinam, Indumathi; Kwasny, Dorota

    2011-01-01

    Microfluidic bioreactors (μBR) are becoming increasingly popular for cell culture, sample preparation and analysis in case of routine genetic and clinical diagnostics. We present a novel μBR for non-adherent cells designed to mimic in vivo perfusion of cells based on diffusion of media through...... a sandwiched membrane. The culture chamber and perfusion chamber are separated by a sandwiched membrane and each chamber has separate inlet/outlets for easy loading/unloading of cells and perfusion of the media. The perfusion of media and exchange of nutrients occur through the sandwiched membrane, which...... was also verified with simulations. Finally, we present the application of this device for cytogenetic sample preparation, whereby we culture and arrest peripheral T-lymphocytes in metaphase and later fix them in the μBR. The expansion of T-lymphocytes from an unknown patient sample was quantified by means...

  8. Meiose analyses of sugar beet triploids

    Directory of Open Access Journals (Sweden)

    Mezei Snežana

    2011-01-01

    Full Text Available During the seed crop growing there can be also problems with uncontrolled pollination if in the previous years there was seed shedding during the harvest, which can cause growth and flowering of diploid, triploid and tetraploid plants and that can present serious problem for the next sugar beet seed crop. Meiosis of triploid hybrids is irregular and can result in development of gametes with different chromosome number. Sugar beet triploids in metaphase I have 9 univalents and 9 bivalents and therefore can be source of aneuploid gametes. After cytological analysis that was performed on 2124 meiocytes, in 14.64% different types of irregularities were detected, such as dislocated chromosomes and chromosome bridges.

  9. Identification of the pattern of heterochromatin distribution in Passiflora species with C-banding.

    Science.gov (United States)

    Viana, A J C; Souza, M M

    2010-09-28

    We tested four C-banding protocols to obtain heterochromatic bands in the passion fruit species Passiflora edulis and P. cacaoensis (Passifloraceae). Three of these protocols had been previously described. The three published protocols were not adequate to obtain C-bands in these species. An adapted protocol demonstrated heterochromatin distribution in metaphasic chromosomes of species of Passiflora for the first time. The differentiated coloration for C-bands was obtained with immersion of the slides in 99% ethanol, 45% acetic acid (additional step), 0.2 N hydrochloric acid, hydroxide of barium, 45% acetic acid, and 2X standard saline citrate at four different temperatures. The C-bands were observed in the satellites and in the telomere and centromere regions of all chromosomes, both in P. edulis and in P. cacaoensis.

  10. Chromosomal radiosensitivity of lymphocytes from Alzheimer's disease patients

    International Nuclear Information System (INIS)

    Tobi, S.E.; Itzhaki, R.F.

    1990-01-01

    We have examined chromosome aberrations in γ irradiated (3 Gy) lymphocytes from five patients with Alzheimer's disease (AD). In each case, the number of dicentrics was significantly higher than the number in irradiated lymphocytes from five age matched normal subjects, the mean value for AD cells being about 25% higher. There was no significant difference in number of acentrics between AD and normal cells. Examination of the number of first, second, and third division metaphases, using fluorescence plus Giemsa staining, indicated that there was no difference in cycling time between AD and normal cells, and that after irradiation both groups showed the same mitotic delay. The similarity of our findings to those of others with irradiated Down's syndrome cells (from adult patients) is discussed. (author)

  11. Distamycin A/DAPI bands and the effects of 5-azacytidine on the chromosomes of the chimpanzee, Pan troglodytes.

    Science.gov (United States)

    Schmid, M; Haaf, T

    1984-01-01

    The chromosomes of the chimpanzee were stained with distamycin A/DAPI, which labels specific C-bands. Bright distamycin A/DAPI fluorescence was found in the heterochromatic regions of chromosomes 6, 11, 14 to 16, 18 to 20, and 23 and the Y. Lymphocyte cultures from chimpanzees were treated with low doses of 5-azacytidine during the last hours of culture. This cytosine analog induces highly distinct undercondensations in 28 heterochromatic regions of 19 chromosomes. These 5-azacytidine-sensitive regions are predominantly located in the terminal C-bands of the chromosomes. In vitro treatment with 5-azacytidine also preserves into the metaphase stage somatic pairings between the 5-azacytidine-sensitive heterochromatic regions in interphase nuclei. The homologies and differences regarding the chromosomal localization of distamycin A/DAPI-bright C-bands, 5-azacytidine-sensitive heterochromatin, 5-methylcytosine-rich DNA sequences, and satellite DNAs in the chimpanzee and man are discussed.

  12. Investigation of relationship between karyotype pattern, effective chromosome-arm number and chromosome aberrations in peripheral blood lymphocytes of three species: man, rabbit, swine. Part of a coordinated programme on radiation induced chromosomal aberrations for genetic risk evaluation in man

    International Nuclear Information System (INIS)

    Liniecki, J.

    1980-08-01

    Blood from 3 donors of each species: man, rabbit and pig was irradiated with a dose of 2.5Gy 60 Co gamma-rays. Microcultures of lymphocytes, established in presence of BrdUrd, were harvested at 6 different times after PHA-stimulation. The preparations containing metaphase figures were stained acc. to Perry and Wolff to permit differentiation of the cells in first and later mitoses. In all individuals and species studied there was a highly significant negative correlation between dicentric yield and time from stimulation to the harvest. The decline of the yield with time of harvest varied in the three species between 1.0 and 3.6 per cent per hour. Implications for biological dosimetry are discussed

  13. Multiple chromosome aberrations among newborns from high level natural radiation area and normal level natural radiation area of south west coast of Kerala

    International Nuclear Information System (INIS)

    Soren, D.C.; Ramachandran, E.N.; Karuppasamy, C.V.; Cheriyan, V.D.; Anil Kumar, V.; Koya, P.K.M.; Seshadri, M.

    2010-01-01

    Cord blood samples were collected in heparin vials and microculture techniques employed to obtain good metaphase chromosome spreads. In cytogenetic studies on newborns cells with multiple aberrations were recorded in 57 from a total of 27285 newborns (1266972 cells). Of these 17294 newborns (964140 cells) were from High Level Natural Radiation Area (HLNRA) and 9991 newborns (302832 cells) from Normal Level Natural Radiation Area (NLNRA). Cells with multiple aberrations were observed in 38 and 19 newborns from High and Normal Level Natural Radiation Area respectively. On an average one cell with multiple aberrations was observed among 479 newborns. Cells with multiple aberrations were observed in newborns from HLNRA as well as NLNRA in both males and females. Gender difference of newborns, maternal age group and background radiation levels did not seem to have any influence in the occurrence of Multiple chromosome aberrations

  14. Application of mammalian cytogenetics to mutagenicity studies

    International Nuclear Information System (INIS)

    Brewen, J.G.

    1977-01-01

    Studies on induction of chromosome damage in germ cells by triethylene melamine (TEM) included determination of frequencies of chromosomal aberrations observed in human leukocytes after treating different stages of the cell cycle with TEM, frequencies of chromatid aberrations in metaphase I oocytes and the female pronuclear chromosomes following treatment of female mice with TEM, and frequencies of labeled diplotene-diakinesis figures and chromosome abberations at various intervals after treatment of primary spermatocytes with TEM and 3 H-thymidine. Studies on effects of low linear energy transfer radiation on mouse oocytes showed that the frequency of aberrations increased as a function of time and remained constant 8 to 9 days post-exposure. It was concluded that cytogenetic procedures were adequate to evaluate certain mutagenic end points

  15. Physical localization of molecular markers and assignment of the 15th linkage group to chromosome 11 of the karyotype in cassava (Manihot esculenta Crantz) by primed in situ labeling.

    Science.gov (United States)

    Wang, Y; Wang, J F; Yin, H; Gao, H Q; Zhuang, N S; Liu, J P

    2015-07-28

    Physical localization of molecular markers and assignment of the 15th linkage group to chromosome 11 of the karyotype in cassava (Manihot esculenta Crantz) were achieved using primed in situ labeling. Amplified signals for both the EST507-1 and SSRY13-5 markers were consistently observed in different stages of cell division. A comparison of the length, arm ratio, and other morphological characteristics of somatic metaphase chromosomes in karyotype analysis indicated that the EST507-1 and SSRY13-5 markers were localized on the short and long arm of cassava chromosome 11 with the relative map positions of 41.67 and 23.07, respectively. The physical localization of the 2 markers on chromosome 11 of the karyotype corresponds to their positions on the 15th linkage group in cassava.

  16. Ultrastructure and mitochondrial numbers in pre- and postpubertal pig oocytes

    DEFF Research Database (Denmark)

    Pedersen, Hanne Skovsgaard; Callesen, Henrik; Løvendahl, Peter

    2016-01-01

    Prepubertal pig oocytes are associated with lower developmental competence. The aim of this experiment was to conduct an exhaustive survey of oocyte ultrastructure and to use a design-unbiased stereological approach to quantify the numerical density and total number of mitochondria in oocytes......, but no differences were observed in mitochondrial densities between groups. Mature postpubertal oocytes adhered to the following characteristics: presence of metaphase II, lack of contact between cumulus cells and oocyte, absence of rough endoplasmic reticulum and Golgi complexes, peripheral location of cortical...... their reduced capacity. The higher number of mitochondria in large pre- and postpubertal oocytes could have an influence on oocyte competence, by increasing the pool of mitochondria available for early embryonic development....

  17. Description of Karyotype of Kerodon acrobata, an endemic rodent in Brazilian Cerrado.

    Science.gov (United States)

    Zappes, I A; Portella, A S; Lessa, G M

    2014-02-01

    Kerodon acrobata is a caviidae rodent endemic from Brazilian Cerrado. It was described only in 1997 and the data about it is very scarce. The aim of this work was to characterize the karyotype of K. acrobata. Giemsa staining, nucleolar organizer region (NOR) banding, C-positive heterochromatin banding and DAPI fluorescence were used in N metaphases of a specimen collected in Asa Branca Farm, in Aurora do Tocantins municipality, Tocantins state, Brazil. K. acrobata showed the same diploid number, fundamental number and chromosome morphology as Kerodon rupestris. But its NOR location and heterochromatin distribution patterns indicated a unique cytogenetic profile when compared to its sister species, emphasizing the evolutionary uniqueness of this relatively new and unknown species. This record also extends the distribution of this species northward.

  18. Towards the theoretical bases of the folding of the 100-A nucleosome filament

    International Nuclear Information System (INIS)

    Chela Flores, J.

    1994-01-01

    We attempt to model DNA packaging at the various stages of ever increasing DNA folding from the 100-A nucleosome filament to various further stages leading up to the metaphase chromosome. We have assumed that a phase transition has induced chromatin into a condensed mode. The mean-field model allows the simultaneous discussion of chromatin with packaging ration η and DNA replication at various stages of folding. We derive a formula correlating (during the S phase of the cell cycle) the DNA polymerase velocity r f (measured in nucleotides per minute) in a relation of inverse proportionality with the degree of DNA packaging: r f = λη -1/2 , where the dimensional constant λ has been determined. This model suggests that in the heterochromatic regions of chromatin there is reduced activity of DNA polymerases. We discuss the possible relevance of our model to late replicating telomeres in yeast and several higher eukaryotes. (author). 28 refs, 3 tabs

  19. The irradiation effects on the cytoskeletons of C3H/He mouse mammary tumor cells and vascular basement membrane in relation to vascular invasion

    International Nuclear Information System (INIS)

    Kobayashi, Michiya

    1988-01-01

    We investigated the short-term effects of a single high-dose radiation upon transplanted MM46 tumor cells in mice by means of immunohistochemistry and electron microscopy. The irradiation induced: 1) giant cell formation from the 3rd day, 2) arrest of tumor cell mitosis in prophase and metaphase due to the disorganization of the mitotic spindles, 3) changes in immunoreactivity of laminin and cytoskeletons, and 4) multilayering of the vascular basal lamina and perivascular fibrosis. The above findings suggest a decrease in tumor cell compliance, growth and invasiveness and the potentiation of defensive host responses against vascular invasion after irradiation. The analysis of the temporal sequences of the events indicates that the time lapse between the optimal host response, tumor growth and invasion constitutes a critical period. (author)

  20. Monitoring the elasticity changes of HeLa cells during mitosis by atomic force microscopy

    Science.gov (United States)

    Jiang, Ningcheng; Wang, Yuhua; Zeng, Jinshu; Ding, Xuemei; Xie, Shusen; Yang, Hongqin

    2016-10-01

    Cell mitosis plays a crucial role in cell life activity, which is one of the important phases in cell division cycle. During the mitosis, the cytoskeleton micro-structure of the cell changed and the biomechanical properties of the cell may vary depending upon different mitosis stages. In this study, the elasticity property of HeLa cells during mitosis was monitored by atomic force microscopy. Also, the actin filaments in different mitosis stages of the cells were observed by confocal imaging. Our results show that the cell in anaphase is stiffer than that in metaphase and telophase. Furthermore, lots of actin filaments gathered in cells' center area in anaphase, which contributes to the rigidity of the cell in this phase. Our findings demonstrate that the nano-biomechanics of living cells could provide a new index for characterizing cell physiological states.

  1. Localization of the tight junction protein gene TJP1 to human chromosome 15q13, distal to the Prader-Willi/Angelman region, and to mouse chromosome 7

    Energy Technology Data Exchange (ETDEWEB)

    Mohandas, T.K. [Darthmouth-Hitchcock Medical Center, Lebanon, NH (United States); Chen, X.N.; Korenberg, J.R. [UCLA School of Medicine, Los Angeles, CA (United States)] [and others

    1995-12-10

    The gene encoding the tight junction (zonula occludens) protein, TJP1, was mapped to human chromosome 15q13 by fluorescence in situ hybridization (FISH) using a cDNA probe. The Jackson Laboratory backcross DNA panel derived from the cross (C57BL/6JEi X SPRET/Ei) F1 females X SPRET/Ei males was used to map the mouse Tjp1 to chromosome 7 near position 30 on the Chromosome Committee Map, a region with conserved homology to human chromosome 15q13. FISH studies on metaphases from patients with the Prader-Willi (PWS) or the Angelman syndrome (AS) showed that TJP1 maps close but distal to the PWS/AS chromosome region. 13 refs., 2 figs.

  2. Parthenogenesis and somatic cell nuclear transfer in sheep oocytes using Polscope.

    Science.gov (United States)

    Nandedkar, Pandit; Chohan, Parul; Patwardhan, Archana; Gaikwad, Santosh; Bhartiya, Deepa

    2009-07-01

    Parthenogenesis and Somatic cell nuclear transfer (SCNT) techniques, offer a unique approach to manipulate the genetic composition of derived human embryonic stem cells - an essential step if the full opportunities for disease modeling, drug discovery or individualized stem cell therapy are to be realized. The present study describes the use of sheep oocytes to acquire expertise and establish methods to reconstruct embryos for obtaining blastocysts before venturing into human SCNT where the oocytes are a very precious starting material. Maturation of sheep eggs in vitro for 20-24 hr resulted in 65% metaphase II (MII) eggs which were either parthenogenetically activated using calcium ionomycin or ethanol or subjected to SCNT using cumulus cell as somatic cell. Sixteen blastocysts were produced by parthenogenetic activation of 350 eggs whereas reconstructed embryos, after SCNT carried out in 139 eggs, progressed only up to morula stage. The procedure of parthenogenesis and SCNT will be useful to generate autologous ES cells using human eggs.

  3. Chromosome-Specific DNA Repeats: Rapid Identification in Silico and Validation Using Fluorescence in Situ Hybridization

    Directory of Open Access Journals (Sweden)

    Heinz-Ulrich G. Weier

    2012-12-01

    Full Text Available Chromosome enumeration in interphase and metaphase cells using fluorescence in situ hybridization (FISH is an established procedure for the rapid and accurate cytogenetic analysis of cell nuclei and polar bodies, the unambiguous gender determination, as well as the definition of tumor-specific signatures. Present bottlenecks in the procedure are a limited number of commercial, non-isotopically labeled probes that can be combined in multiplex FISH assays and the relatively high price and effort to develop additional probes. We describe a streamlined approach for rapid probe definition, synthesis and validation, which is based on the analysis of publicly available DNA sequence information, also known as “database mining”. Examples of probe preparation for the human gonosomes and chromosome 16 as a selected autosome outline the probe selection strategy, define a timeline for expedited probe production and compare this novel selection strategy to more conventional probe cloning protocols.

  4. Cell-cycle calcium transients driven by cyclic changes in inositol trisphosphate levels.

    Science.gov (United States)

    Ciapa, B; Pesando, D; Wilding, M; Whitaker, M

    1994-04-28

    Transient changes in intracellular calcium ([Ca2+]i) have been shown to punctuate the cell cycle in various types of cells in culture and in early embryos. The [Ca2+]i transients are correlated with cell-cycle events: pronuclear migration, nuclear envelope breakdown, the metaphase-anaphase transition of mitosis, and cytokinesis. Mitotic events can be induced by injecting calcium and prevented by injecting calcium chelators into the sea urchin embryo. Cell-cycle calcium transients differ from the transients linked to membrane signal transduction pathways: they are generated by an endogenous mechanism, not by plasma membrane receptor complexes, and their trigger is unknown. We report here that the phosphoinositide messenger system oscillates during the early embryonic cell cycle in the sea urchin, leading to cyclic increases in inositol trisphosphate that trigger cell-cycle [Ca2+]i transients and mitosis by calcium release from intracellular stores.

  5. Protective effect of vitamins C and E on Gamma radiation induced Genetic injuries in male mice germ cells

    International Nuclear Information System (INIS)

    Anwar, W.A.; El-Daway, H.A.E.; Tawfik, S.S.M.

    1999-01-01

    The effects of vitamins C and E on meiotic chromosomal metaphase-8 at diakinesis of the mouse to 3 Gy of whole body gamma- irradiation were studied. These vitamins were injected intraperitoneally as acute doses 2 hr before irradiation. Both vitamins significantly reduced the frequencies of chromosomal aberration in spermatic germ cells. The protective effect of vitamin E was greater than that afforded by vitamin C. A combined treatment of both vitamins resulted in additional protection over that offered by each vitamine alone. In all animal groups the most frequent aberration found was translocation in the from of either ring four (R IV) or chain four (C IV). The percentage of each or them was significantly increased in male mice sacrificed after 15 days post-irradiation. Other types of aberrations as autosomal univalent, X-Gamma univalent and polyploidy were rarely present

  6. RCC1 regulates inner centromeric composition in a Ran-independent fashion.

    Science.gov (United States)

    Zhang, Michael Shaofei; Furuta, Maiko; Arnaoutov, Alexei; Dasso, Mary

    2018-04-05

    RCC1 associates to chromatin dynamically within mitosis and catalyzes Ran-GTP production. Exogenous RCC1 disrupts kinetochore structure in Xenopus egg extracts (XEEs), but the molecular basis of this disruption remains unknown. We have investigated this question, utilizing replicated chromosomes that possess paired sister kinetochores. We find that exogenous RCC1 evicts a specific subset of inner KT proteins including Shugoshin-1 (Sgo1) and the chromosome passenger complex (CPC). We generated RCC1 mutants that separate its enzymatic activity and chromatin binding. Strikingly, Sgo1 and CPC eviction depended only on RCC1's chromatin affinity but not its capacity to produce Ran-GTP. RCC1 similarly released Sgo1 and CPC from synthetic kinetochores assembled on CENP-A nucleosome arrays. Together, our findings indicate RCC1 regulates kinetochores at the metaphase-anaphase transition through Ran-GTP-independent displacement of Sgo1 and CPC.

  7. Partial trisomy 13q identified by sequential fluorescence in situ hybridization

    Energy Technology Data Exchange (ETDEWEB)

    Gopal Rao, V.V.N.; Carpenter, N.J.; Gucsavas, M. [Institute of Medical Genetics, Tulsa, OK (United States)] [and others

    1995-07-31

    We report on a 19-month-old boy with partial trisomy 13q resulting from a probable balanced translocation involving chromosomes 1 and 13. The infant presented with omphalocele, malrotation, microcephaly with overriding skull bones, micrognathia, apparently low-set ears, rocker-bottom feet, and congenital heart disease, findings suggestive of trisomy 13. Karyotypic studies from peripheral blood lymphocytes documented an unbalanced karyotype 46,XY,-1,+der. The mother`s chromosomes were normal, and the father was not available. Conventional cytogenetic techniques were unable to identify the extra material on the terminal 1q. Using fluorescence in situ hybridization (FISH) on the GTL-banded metaphases, the extra material on 1q was identified as the terminal long arm of 13, thus resulting in partial trisomy 13 (q32-qter). 8 refs., 2 figs., 1 tab.

  8. Assessment of DNA Damage in Peripheral Blood Lymphocytes of Radiation Workers at Al-Tuwaitha Site by Using the Sister Chromatid Exchange and the Comet Assay

    International Nuclear Information System (INIS)

    Ali, A.K.; Muttar, A.J.; Khayon, S.K.; Haider, Y.L.; Ali, H.F.; Abdullah, A.K.

    2015-01-01

    The sister chromatid exchange was performed on peripheral blood lymphocytes obtained from 40 individuals of workers occupationally exposed to low ionizing radiation doses in Al-Tuwaitha site due to decommissioning to radioactive contamination then compared with 40 control individuals living in Baghdad. SCEs were scored in metaphase chromosomes were identified by fluorescent plus Giemsa staining (Figure 2).The mean frequencies of SCEs per cell differed significantly (p≺0 0.05) between individuals of radiation workers and control, being 7.78 0.45 SCE/cells and 6.28 0.22 SCE/cells , respectively. However SCE frequency was statistically significant (P≺0 0.05) among radiation workers as compared to control individuals.

  9. Influence of Low-Dose Ionising Radiation on the Occurrence of Chromosomal Aberrations in Horse Lymphocytes after In vitro Irradiation

    International Nuclear Information System (INIS)

    Rukavina, D.; Hasanbasic, D.; Haveric, A.; Haveric, S.; Ajanovic, A.; Katica, A.

    2008-01-01

    It is well known that ionising radiation affect the chromosomal aberrations that have been used as a reliable criteria in biological dosimetry. Many studies confirm that the increase of radiation doses is related to the increase of mutation number and that even low doses could induce the mutations. Thus in this study we analysed the influence of low-dose ionising radiation to the occurrence of chromosomal aberrations in lymphocytes of peripheral blood of 12 horses after in vitro X-irradiation. To gain metaphase plates the method of cultivation periphery blood lymphocytes was employed. The observed aberrations were recorded and classified according to the International System of Cytogenetic Nomenclature (ISCN). The results showed that the percentage of chromosomal aberrations in irradiated blood (0.1 Gy and 0.2 Gy) was higher relative to the controlled samples of nonirradiated blood.(author)

  10. Chromosomal damage and mitotic index in Bosnian and Herzegovinian mountain horse lymphocytes following low dosage X-ray irradiation in vitro

    International Nuclear Information System (INIS)

    Rukavina, D.; Hasanbašić, D.; Katica, A.; Suljkanović, E.

    2009-01-01

    In this study we investigated the influence of low dosage X-ray irradiation on the incidence of chromosomal damage and changes in mitotic index (MI) in cultured peripheral lymphocytes of Bosnian and Herzegovinian mountain horse following in vitro irradiation. X-ray irradiation induced a dose-dependent decrease in MI but only the dose of 0.5 Gy induced a significant decrease in comparison with the control and other dose groups. The analysis the chromosomal damage revealed a clear dose-dependent increase in the incidence of chromosomal damage per metaphase. Significant differences were detected by analysis of variance and the LSD test confirmed significant differences between cell that received 0.2 Gy and 0.5 Gy when compared to the control cells and cells that received 0.1 Gy. However, Scheeffes' test assigned significance only to the differences established between the cells that received 0.5 Gy and the other groups of lymphocytes

  11. Molecular cloning and chromosome mapping of the human gene encoding protein phosphotyrosyl phosphatase 1B

    International Nuclear Information System (INIS)

    Brown-Shimer, S.; Johnson, K.A.; Bruskin, A.; Green, N.R.; Hill, D.E.; Lawrence, J.B.; Johnson, C.

    1990-01-01

    The inactivation of growth suppressor genes appears to play a major role in the malignant process. To assess whether protein phosphotyrosyl phosphatases function as growth suppressors, the authors have isolated a cDNA clone encoding human protein phosphotyrosyl phosphatase 1B for structural and functional characterization. The translation product deduced from the 1,305-nucleotide open reading frame predicts a protein containing 435 amino acids and having a molecular mass of 49,966 Da. The amino-terminal 321 amino acids deduced from the cDNA sequence are identical to the empirically determined sequence of protein phosphotyrosyl phosphatase 1B. A genomic clone has been isolated and used in an in situ hybridization to banded metaphase chromosomes to determine that the gene encoding protein phosphotyrosyl phosphatase 1B maps as a single-copy gene to the long arm of chromosome 20 in the region q13.1-q13.2

  12. Fluorescence cell imaging and manipulation using conventional halogen lamp microscopy.

    Directory of Open Access Journals (Sweden)

    Kazuo Yamagata

    Full Text Available Technologies for vitally labeling cells with fluorescent dyes have advanced remarkably. However, to excite fluorescent dyes currently requires powerful illumination, which can cause phototoxic damage to the cells and increases the cost of microscopy. We have developed a filter system to excite fluorescent dyes using a conventional transmission microscope equipped with a halogen lamp. This method allows us to observe previously invisible cell organelles, such as the metaphase spindle of oocytes, without causing phototoxicity. Cells remain healthy even after intensive manipulation under fluorescence observation, such as during bovine, porcine and mouse somatic cell cloning using nuclear transfer. This method does not require expensive epifluorescence equipment and so could help to reduce the science gap between developed and developing countries.

  13. Mouse Mos protooncogene product is present and functions during oogenesis.

    Science.gov (United States)

    Paules, R S; Buccione, R; Moschel, R C; Vande Woude, G F; Eppig, J J

    1989-07-01

    We have identified the mouse Mos-encoded protein product, p39mos, in maturing mouse oocytes and have shown that it is indistinguishable from the product expressed in Mos-transformed NIH 3T3 cells. p39mos is detected in oocytes arrested in the first meiotic prophase, during germinal-vesicle breakdown, metaphase I, anaphase I, and in ovulated eggs. We show that microinjection of three different Mos antisense (but not sense) oligodeoxyribonucleotides into germinal vesicle-stage oocytes prevents first polar-body emission and therefore interrupted the normal progression of meiosis. These results show that in mouse oocytes, as in the amphibian Xenopus [Sagata, N., Oskarsson, M., Copeland, T., Brumbaugh, J. & Vande Woude, G.F. (1988) Nature (London) 335, 519-525], the product of Mos is necessary for normal meiotic maturation.

  14. Studies on radiation-induced chromosome damage in humans: Semi-annual progress report, October 1, 1986-March 31, 1987

    International Nuclear Information System (INIS)

    Littlefield, L.G.

    1987-01-01

    This report summarizes recent research to determine and report the frequency of somatic cell chromosome aberrations in approximately 200 lymphocyte metaphases from each of 200 control patients or persons who received radiation for enlarged thymus, and from an additional 475 irradiated and control subjects selected by NCI from populations exposed to therapeutic ionizing radiation during the period 1930 to 1970. The priority of populations to be studied will be determined by NCI in consultation with the contractor and with advice from NCI consultants. Additional research will determine and report dose response curves among the several populations, to determine how differences with respect to radiation dose, quality of radiation, fractionation, sex and age within and among groups affect the ''dose-response relationship.'' 7 tabs

  15. Chromosome investigations on persons whose mothers have been treated with ionizing radiations during pregnancy

    International Nuclear Information System (INIS)

    Hanebuth, P.

    1982-01-01

    This thesis reports on chromosome investigations on seven persons between 2 and 26 years of age who, during their prenatal life, have been exposed to ionizing radiation. The metaphase chromosomes from peripheral lymphocytes, obtained by standard cytogenic methods, have been scanned for numerical and structural aberrations after a culture period of two or three days. Comparison with non-irradiated specimens revealed a summarily slight increase in the occurrence of some structural aberrations in the irradiated material. The differences are smaller in number than the rate of deviation to be accounted to culturing techniques so that one cannot deduce a radiation-induced impairment from these results. As one of the persons investigated has been undergoing longterm therapy with anticonvulsive drugs, this case is discussed separately. In another case, new staining methods (G, C, or Q-bands) together with an investigation of material from the father, revealed a particularly small Y-chromosome. (orig./MG) [de

  16. Syncytes with premeiotic mitotic and cytomictic comportment in opium poppy (Papaver somniferum L.)

    International Nuclear Information System (INIS)

    Patra, N.K.; Chauhan, S.P.; Srivastava, H.K.

    1987-01-01

    Variations in terms of mitosis at premeiotic stage and cytomixis in multiploid microsporocytes have been recorded in mutagen treated and untreated populations raised from an inbred line S4I30 of Papaver sominiferum L. (2n = 22). In premeiotic mitosis all the chromosomes in syncytes were found to align on a single metaphase plate and separate normally in anaphase. Ploidy levels in premeiotic syncytes varied considerably from 2N to 4N in control, from 2N to 6N in 5kR-M1 and 2N to 10N in combined dose (5kR + 0.6% EMS) M1. Specific heterochromatic chromosomes were observed to be involved in cytomictic events suggesting thereby that cytomixis is a genetically manoeuvred process for the generation of syncytes. The results have been discussed from the point of view that higher ploidy level coupled with less sterility in gametes may be instrumental for variation and evolution in opium poppy

  17. The human insulin receptor substrate-1 gene (IRS1) is localized on 2q36

    Energy Technology Data Exchange (ETDEWEB)

    Nishiyama, Masaki; Matsufuji, Senya; Hayashi, Shin-ichi; Furusaka, Akihiro; Tanaka, Teruji (Jikei Univ. School of Medicine, Tokyo (Japan)); Inazawa, J.; Nakamura, Yusuke (Cancer Institute, Tokyo (Japan)); Ariyama, Takeshi (Kyoto Prefactural Univ. of Medicine (Japan)); Wands, J.R. (Harvard Medical School, Boston, MA (United States))

    1994-03-01

    The chromosomal localization of some of the genes participating in the insulin signaling pathway is known. The insulin and insulin receptor genes have been mapped to chromosomes 11 and 19, respectively. To identify the chromosomal localization of the human IRS1 gene, the fluorescence in situ hybridization technique was employed with Genomic Clone B-10. A total of 50 metaphase cells exhibiting either single or double spots of hybridization signals were examined. Among them, 32 showed the specific signals on 2q36. Therefore, the authors assigned the human IRS1 gene to 2q36. The genes for homeobox sequence (HOX4), fibronectin 1, alkaline phosphatase (intestinal), transition protein 1, villin 1, collagen (type IV), Waardenburg syndrome (type 1), alanine-glyoxylate aminotransferase, and glucagon have been localized in the vicinity of the IRS1 gene.

  18. Cell inactivation and chromosomal aberrations induced by X-rays and fast neutrons in cells of the Chinese hamster. 1

    International Nuclear Information System (INIS)

    Tolkendorf, E.

    1979-01-01

    Asynchronously grown cultures of Chinese hamster cells V79-4 were irradiated in suspension with 180 kV X-rays and fast neutrons (average energy of 6.2 MeV). The damage was assessed by measuring cell survival and frequencies of chromosome aberrations in the first post-irradiation metaphases. The experimental data for survival and chromosome aberrations were fitted by computer programmes. From the fitted curves the relative biological effectiveness (RBE) of fast neutrons was calculated. The RBE shows a similar dose dependence for killed and aberrant cells. The RBE decreases with increasing dose and amounts to approximately 5 for both effects for small neutron doses. The highest RBE is found for asymmetrical chromosomal exchanges and is dependent on the neutron dose, too. However, for isochromatid deletions the RBE is dose independent with a value of 3.6. (author)

  19. Chromosome characterization of two varieties of Mangifera indica L.¹

    Directory of Open Access Journals (Sweden)

    Neiva Izabel Pierozzi

    2011-10-01

    Full Text Available Chromosome studies were performed in two varieties of Mangifera indica L. (mango, 'IAC-140 Espadona' and in its progenitor 'Espada Stahl'. Both varieties showed 2n=40 chromosomes though the karyotype formulae were 8m + 10sm + 2sm s for 'Stahl' and 7m + 11sm + 2sm s for 'IAC-140'. The varieties showed moderate karyotype asymmetry which was estimated according to four different indices. Both varieties exhibited three chromosome pairs with silver impregnation after NOR-banding. The number of nucleoli within interphase cells varied from one, the commonest, to eight. The nucleolus persistent phenomenon was observed in more than 22% of metaphase cells of both varieties, seeing that in 'Stahl', up to two nucleoli were evidenced. This variety also showed one nucleolus in several anaphase cells. The studies were suitable for evidencing diversity at chromosomal level between these two varieties.

  20. Intercentrosomal angular separation during mitosis plays a crucial role for maintaining spindle stability

    Science.gov (United States)

    Sutradhar, S.; Basu, S.; Paul, R.

    2015-10-01

    Cell division through proper spindle formation is one of the key puzzles in cell biology. In most mammalian cells, chromosomes spontaneously arrange to achieve a stable bipolar spindle during metaphase which eventually ensures proper segregation of the DNA into the daughter cells. In this paper, we present a robust three-dimensional mechanistic model to investigate the formation and maintenance of a bipolar mitotic spindle in mammalian cells under different physiological constraints. Using realistic parameters, we test spindle viability by measuring the spindle length and studying the chromosomal configuration. The model strikingly predicts a feature of the spindle instability arising from the insufficient intercentrosomal angular separation and impaired sliding of the interpolar microtubules. In addition, our model successfully reproduces chromosomal patterns observed in mammalian cells, when activity of different motor proteins is perturbed.

  1. Aurora-A regulates MCRS1 function during mitosis.

    Science.gov (United States)

    Meunier, Sylvain; Timón, Krystal; Vernos, Isabelle

    2016-07-02

    The mitotic spindle is made of microtubules (MTs) nucleated through different pathways involving the centrosomes, the chromosomes or the walls of pre-existing MTs. MCRS1 is a RanGTP target that specifically associates with the chromosome-driven MTs protecting them from MT depolymerases. MCRS1 is also needed for the control of kinetochore fiber (K-fiber) MT minus-ends dynamics in metaphase. Here, we investigated the regulation of MCRS1 activity in M-phase. We show that MCRS1 is phosphorylated by the Aurora-A kinase in mitosis on Ser35/36. Although this phosphorylation has no role on MCRS1 localization to chromosomal MTs and K-fiber minus-ends, we show that it regulates MCRS1 activity in mitosis. We conclude that Aurora-A activity is particularly important in the tuning of K-fiber minus-ends dynamics in mitosis.

  2. Cell cycle regulation of human immunodeficiency virus type 1 integration in T cells: antagonistic effects of nuclear envelope breakdown and chromatin condensation

    International Nuclear Information System (INIS)

    Mannioui, Abdelkrim; Schiffer, Cecile; Felix, Nathalie

    2004-01-01

    We examined the influence of mitosis on the kinetics of human immunodeficiency virus type 1 integration in T cells. Single-round infection of cells arrested in G1b or allowed to synchronously proceed through division showed that mitosis delays virus integration until 18-24 h postinfection, whereas integration reaches maximum levels by 15 h in G1b-arrested cells. Subcellular fractionation of metaphase-arrested cells indicated that, while nuclear envelope disassembly facilitates docking of viral DNA to chromatin, chromosome condensation directly antagonizes and therefore delays integration. As a result of the balance between the two effects, virus integration efficiency is eventually up to threefold greater in dividing cells. At the single-cell level, using a green fluorescent protein-expressing reporter virus, we found that passage through mitosis leads to prominent asymmetric segregation of the viral genome in daughter cells without interfering with provirus expression

  3. Mosaic 22q11.2 microdeletion syndrome: diagnosis and clinical manifestations of two cases

    Directory of Open Access Journals (Sweden)

    Kabra Madhulika

    2008-08-01

    Full Text Available Abstract Chromosome 22q11.2 microdeletion syndrome is due to microdeletion of 22q11.2 region of chromosome 22. It is a common microdeletion syndrome however mosaic cases are very rare and reported only few previous occasions. In this report we describe two unrelated male children with clinical features consistent with 22q11.2 microdeletion syndrome characterized by cardiac defect, facial dysmorphism and developmental deficiency. One of the cases also had trigonocephaly. Interphase & metaphase FISH with 22q11.2 probe demonstrated mosaicism for hemizygous deletion of 22q11.2 region. Mosaicism is also observed in buccal cells as well as urine cells. Parents were without any deletion. These two cases represent rare cases of mosaic 22q11.2 microdeletion syndrome.

  4. Mosaic 22q11.2 microdeletion syndrome: diagnosis and clinical manifestations of two cases.

    Science.gov (United States)

    Halder, Ashutosh; Jain, Manish; Kabra, Madhulika; Gupta, Neerja

    2008-08-10

    Chromosome 22q11.2 microdeletion syndrome is due to microdeletion of 22q11.2 region of chromosome 22. It is a common microdeletion syndrome however mosaic cases are very rare and reported only few previous occasions. In this report we describe two unrelated male children with clinical features consistent with 22q11.2 microdeletion syndrome characterized by cardiac defect, facial dysmorphism and developmental deficiency. One of the cases also had trigonocephaly. Interphase & metaphase FISH with 22q11.2 probe demonstrated mosaicism for hemizygous deletion of 22q11.2 region. Mosaicism is also observed in buccal cells as well as urine cells. Parents were without any deletion. These two cases represent rare cases of mosaic 22q11.2 microdeletion syndrome.

  5. The need for additional genetic markers for MDS stratification: what does the future hold for prognostication?

    Science.gov (United States)

    Otrock, Zaher K.; Tiu, Ramon V.; Maciejewski, Jaroslaw P.; Sekeres, Mikkael A.

    2013-01-01

    Myelodysplastic syndromes (MDS) constitute a heterogeneous group of clonal hematopoietic disorders. Metaphase cytogenetics (MC) has been the gold standard for genetic testing in MDS, but it can detect clonal cytogenetic abnormalities in only 50% of cases. New karyotyping tests include fluorescence in situ hybridization (FISH), array-based comparative genomic hybridization (aCGH), and single nucleotide polymorphism arrays (SNP-A). These techniques have increased the detected genetic abnormalities in MDS, many of which confer prognostic significance to overall and leukemia-free survival. This has eventually increased our understanding of MDS genetics. With the help of new technologies, we anticipate that the existing prognostic scoring systems will incorporate mutational data into their parameters. This review discusses the progress in MDS diagnosis through the use of array-based technologies. We also discuss the recently investigated genetic mutation in MDS, and revisit the MDS classification and prognostic scoring systems. PMID:23373781

  6. Postnatal diagnosis of constitutive ring chromosome 13 using both conventional and molecular cytogenetic approaches.

    Science.gov (United States)

    Minasi, L B; Pinto, I P; de Almeida, J G; de Melo, A V; Cunha, D M C; Ribeiro, C L; Silva, G P; Brasil, M G; Silva, D M; da Silva, C C; da Cruz, A D

    2015-03-06

    We describe the first postnatal diagnosis of a child from Central Brazil with de novo cytogenetic alterations in 13q showing malformations of the brain, eyes, distal limbs, and genitourinary tract, and severe intellectual disability. The karyotype was a constitutive 46,XX,r(13)[77]/45,XX,-13[17]/46,XX,idic r(13)[6]. Interphase and metaphase fluorescence in situ hybridization analyses also showed the absence of 13qter and the presence of 13q14.3 in the cells with r(13), and chromosome microarray analysis detected a 15.39 Mb deletion in chromosome region 13q32.3-q34. This study is intended as the registry of a rare case of chromosomal rearrangement involving chromosome 13 in Central Brazil. Further studies are needed to define whether genetic haploinsufficiency is associated with each major 13q deletion anomaly.

  7. Nucleolar organizer regions in Sittasomus griseicapillus and Lepidocolaptes angustirostris (Aves, Dendrocolaptidae: evidence of a chromosome inversion

    Directory of Open Access Journals (Sweden)

    Marcelo de Oliveira Barbosa

    2013-01-01

    Full Text Available Cytogenetic studies in birds are still scarce compared to other vertebrates. Woodcreepers (Dendrocolaptidae are part of a highly specialized group within the Suboscines of the New World. They are forest birds exclusive to the Neotropical region and similar to woodpeckers, at a comparable evolutionary stage. This paper describes for the first time the karyotypes of the Olivaceous and the Narrow-billed Woodcreeper using conventional staining with Giemsa and silver nitrate staining of the nucleolar organizer regions (Ag-NORs. Metaphases were obtained by fibular bone marrow culture. The chromosome number of the Olivaceous Woodcreeper was 2n = 82 and of the Narrow-billed Woodcreeper, 2n = 82. Ag-NORs in the largest macrochromosome pair and evidence of a chromosome inversion are described herein for the first time for this group.

  8. The neuroblast of the grasshopper embryo as a new mutagen test system. Pt. 1

    International Nuclear Information System (INIS)

    Liang, J.C.; Gaulden, M.E.

    1982-01-01

    The neuroblasts of the grasshopper embryo (Chortophaga viridifasciata De Geer) are being studied to determine their suitability for detecting environmental clastogens (chromosome-breaking agents). They are very sensitive to the induction of chromosome breakage by radiation in viro. Their sensitvity, 0.011 break/cell/R, is 4-5 times higher than pollen mother cells of Tradescantia (micronuclei), 10 times higher than either human lymphocytes or Chinese hamster cells (metaphase chromosome aberrations), and 15 times higher than mouse erythroblasts (micronuclei). Furthermore, they have no spontaneous chromosome breakage, which facilitates the detection of agents that break chromosomes. The present study shows that Chortophaga embryos maintain normal mitotic activity in vitro for 5 cell cycles at 38 0 C (20 h), and that neuroblasts of embryos grown in vitro have the same radiosensitivity as those of embryos in vivo. Thus in vitro exposure of grasshopper embryos is a promising method for obtaining data on the response of neuroblasts to chemical clastogens. (orig.)

  9. Chromosome-specific staining to detect genetic rearrangements associated with chromosome 3 and/or chromosome 17

    Energy Technology Data Exchange (ETDEWEB)

    Gray, Joe W [San Francisco, CA; Pinkel, Daniel [Lafayette, CA; Kallioniemi, Olli-Pekka [Turku, FI; Kallioniemi, Anne [Tampere, FI; Sakamoto, Masaru [Tokyo, JP

    2009-10-06

    Methods and compositions for staining based upon nucleic acid sequence that employ .[.nudeic.]. .Iadd.nucleic .Iaddend.acid probes are provided. Said methods produce staining patterns that can be tailored for specific cytogenetic analyses. Said probes are appropriate for in situ hybridization and stain both interphase and metaphase chromosomal material with reliable signals. The nucleic acid probes are typically of a complexity greater than 50 kb, the complexity depending upon the cytogenetic application. Methods and reagents are provided for the detection of genetic rearrangements. Probes and test kits are provided for use in detecting genetic rearrangements, particularly for use in tumor cytogenetics, in the detection of disease related loci, specifically cancer, such as chronic myelogenous leukemia (CML), retinoblastoma, ovarian and uterine cancers, and for biological dosimetry. Methods and reagents are described for cytogenetic research, for the differentiation of cytogenetically similar but genetically different diseases, and for many prognostic and diagnostic applications.

  10. Image processing in biodosimetry: A proposal of a generic free software platform.

    Science.gov (United States)

    Dumpelmann, Matthias; Cadena da Matta, Mariel; Pereira de Lemos Pinto, Marcela Maria; de Salazar E Fernandes, Thiago; Borges da Silva, Edvane; Amaral, Ademir

    2015-08-01

    The scoring of chromosome aberrations is the most reliable biological method for evaluating individual exposure to ionizing radiation. However, microscopic analyses of chromosome human metaphases, generally employed to identify aberrations mainly dicentrics (chromosome with two centromeres), is a laborious task. This method is time consuming and its application in biological dosimetry would be almost impossible in case of a large scale radiation incidents. In this project, a generic software was enhanced for automatic chromosome image processing from a framework originally developed for the Framework V project Simbio, of the European Union for applications in the area of source localization from electroencephalographic signals. The platforms capability is demonstrated by a study comparing automatic segmentation strategies of chromosomes from microscopic images.

  11. Isochromosome 9q as a sole anomaly in an Omani boy with acute lymphoblastic leukaemia

    Science.gov (United States)

    Achandira, Udayakumar Muthappa; Pathare, Anil V; Kindi, Salam Al; Dennison, David; Yahyaee, Said Al

    2009-01-01

    This report describes a case of acute lymphoblastic leukaemia in which isochromosome 9q (i(9q)) was the sole acquired cytogenetic abnormality. The Immunophenotype showed positivity for CD3, CD4, CD5, CD7, CD8, CD10, CD71, CD117 and TdT, consistent with T cell acute lymphoblastic leukaemia (ALL). The chromosomal analysis of bone marrow showed 46,XY,i(9)(q10) in all the metaphases analysed. The bone marrow morphology was ALL-L2 as per the French–American–British criteria. Isochromosomes are rare chromosomal abnormalities in childhood ALL and the effect of i(9q) is not well established. The patient’s good response to therapy with normal cytogenetics within a month of induction, and disease-free survival after bone marrow transplant are indicative of a good prognosis in such cases. PMID:21686579

  12. The brain finger protein gene (ZNF179), a member of the RING finger family, maps within the Smith-Magenis syndrome region at 17p11.2

    Energy Technology Data Exchange (ETDEWEB)

    Kimura, Toshiyuki; Arakawa, Yoshiki; Inazawa, Johji [Kyoto Prefectural Univ. of Medicine, Kyoto (Japan)] [and others

    1997-03-31

    Smith-Magenis syndrome (SAIS) is caused by a microdeletion of 17p11.2 and comprises developmental and growth delay, facial abnormalities, unusual behavior and sleep problems. This phenotype may be due to haploinsufficiency of several contiguous genes. The human brain finger protein gene (ZNF179), a member of the RING finger protein family, has been isolated and mapped to l7p11.2. FISH analyses of metaphase or interphase chromosomes of 6 patients with SMS show that ZNF179 was deleted in one of the 2 homologs (17p11.2), indicating a possible association of the defect of this gene with the pathogenesis of SMS. Furthermore, using a prophase FISH ordering system, we sublocalized ZNF179 proximally to LLGL which lies on the critical region for SMS. 27 refs., 2 figs.

  13. Method of detecting genetic deletions identified with chromosomal abnormalities

    Energy Technology Data Exchange (ETDEWEB)

    Gray, Joe W; Pinkel, Daniel; Tkachuk, Douglas

    2013-11-26

    Methods and compositions for staining based upon nucleic acid sequence that employ nucleic acid probes are provided. Said methods produce staining patterns that can be tailored for specific cytogenetic analyzes. Said probes are appropriate for in situ hybridization and stain both interphase and metaphase chromosomal material with reliable signals. The nucleic acids probes are typically of a complexity greater tha 50 kb, the complexity depending upon the cytogenetic application. Methods and reagents are provided for the detection of genetic rearrangements. Probes and test kits are provided for use in detecting genetic rearrangements, particlularly for use in tumor cytogenetics, in the detection of disease related loci, specifically cancer, such as chronic myelogenous leukemia (CML) and for biological dosimetry. Methods and reagents are described for cytogenetic research, for the differentiation of cytogenetically similar ut genetically different diseases, and for many prognostic and diagnostic applications.

  14. Chromosome-specific staining to detect genetic rearrangements

    Energy Technology Data Exchange (ETDEWEB)

    Gray, Joe W.; Pinkel, Daniel; Tkachuk, Douglas; Westbrook, Carol

    2013-04-09

    Methods and compositions for staining based upon nucleic acid sequence that employ nucleic acid probes are provided. Said methods produce staining patterns that can be tailored for specific cytogenetic analyzes. Said probes are appropriate for in situ hybridization and stain both interphase and metaphase chromosomal material with reliable signals. The nucleic acid probes are typically of a complexity greater than 50 kb, the complexity depending upon the cytogenetic application. Methods and reagents are provided for the detection of genetic rearrangements. Probes and test kits are provided for use in detecting genetic rearrangements, particularly for use in tumor cytogenetics, in the detection of disease related loci, specifically cancer, such as chronic myelogenous leukemia (CML) and for biological dosimetry. Methods and reagents are described for cytogenetic research, for the differentiation of cytogenetically similar but genetically different diseases, and for many prognostic and diagnostic applications.

  15. Origin of meiotic nondisjunction in Drosophila females

    International Nuclear Information System (INIS)

    Grell, R.F.

    1978-01-01

    Meiotic nondisjunction can be induced by external agents, such as heat, radiation, and chemicals, and by internal genotypic alterations, namely, point mutations and chromosomal rearrangements. In many cases nondisjunction arises from a reduction or elimination of crossing-over, leading to the production of homologous univalents which fail to co-orient on the metaphase plate and to disjoin properly. In some organisms, e.g., Drosophila and perhaps man, distributive pairing [i.e., a post-exchange, size-dependent pairing] ensures the regular segregation of such homologous univalents. When a nonhomologous univalent is present, which falls within a size range permitting nonhomologous recognition and pairing, distributive nondisjunction of the homologues may follow. Examples of nondisjunction induced by inversion heterozygosity, translocation heterozygosity, chromosome fragments, radiation, heat, and recombination-defective mutants are presented

  16. A study of the first and second polar bodies in mouse oogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Evsikov, A.V.; Evsikov, S.V. [Institute of Molecular Biology and Genetics, Kiev (Ukraine)

    1995-05-01

    The possibility of using a polar body for biopsy of human oocytes and early embryos has recently been shown. Genetic analysis of polar bodies can also provide additional information about the mechanisms underlying oocyte maturation. The first polar body is extremely unstable: in mouse oocytes, it disintegrates within several hours. Thus, the possibilities for its analysis are limited. We obtained karyoplasts of mouse eggs that contained the metaphase II spindle. By using them as a model for the first polar body, we studied the causes of its rapid disintegration. The rates of disintegration of the karyoplasts treated with various inhibitors of the cytoskeleton indicate that disintegration of the first polar body may be due to interaction between the actin cytoskeleton, chromatin, and the plasma membrane. The second polar body is found in mouse embryos until the blastocyst stage. Fusion of the second polar body with the enucleated zygote allowed analysis of its chromosomes. 22 refs., 5 figs., 1 tab.

  17. Delayed chromosomal instability induced by DNA damage

    International Nuclear Information System (INIS)

    Morgan, W.F.; Marder, B.A.; Day, J.P.

    1994-01-01

    Cellular exposure to DNA damaging agents rapidly results in a dose dependent increase in chromosomal breakage and gross structural chromosomal rearrangements. Over recent years, evidence has been accumulating indicating genomic instability can manifest multiple generations after cellular exposure to physical and chemical DNA damaging agents. Genomic instability manifests in the progeny of surviving cells, and has been implicated in mutation, gene application, cellular transformation, and cell killing. To investigate chromosome instability following DNA damage, we have used fluorescence in situ hybridization to detect chromosomal rearrangements in a human/hamster somatic hybrid cell line following exposure to ionizing radiation. Delayed chromosomal instability was detected when multiple populations of uniquely arranged metaphases were observed in clonal isolates raised from single cells surviving X-irradiation many generations after exposure. At higher radiation doses, chromosomal instability was observed in a relatively high frequency of surviving clones and, in general, those clones showed delayed chromosome instability also showed reduced survival as measured by colony forming ability

  18. Genetic and cytogenetic characterization of genetic sexing strains of Bactrocera dorsalis and Bactrocera cucurbitae (Diptera: Tephritidae).

    Science.gov (United States)

    Zacharopoulou, A; Franz, G

    2013-04-01

    In the current study, we performed genetic and cytogenetic analyses of two genetic sexing strains (GSSs), one for Bactrocera dorsalis s.s. and one for melon fly, Bactrocera cucurbitae Coquillett, the first such strains ever constructed in these species. In both strains, the genetic sexing mechanism is based on a pupal color dimorphism (white or brown) and is the result of a reciprocal translocation between the Y chromosome and the autosome bearing the white pupae (wp) locus. Based on genetic analysis and cytological data on mitotic metaphases and larval salivary gland polytene chromosomes, we succeeded in mapping the autosome breakpoints in the two Y-autosome translocations even though the Y chromosome is not visible in polytene nuclei. We show that polytene chromosomes can be used in cytogenetic analyses toward the development of genetic control methods in these pest species. The results of the genetic analysis are in full agreement with the cytological description of the strains.

  19. Cytogenetics of Triatominae: III - A study on male sterility induced through hybridization of Triatoma sórdida and Triatoma pseudomaculata

    Directory of Open Access Journals (Sweden)

    Giorgio Schreiber

    1975-08-01

    Full Text Available Males from bilateral crosses between Triatoma sórdida and Triatoma pseudomaculata were unable to give offspring, as shown by subsequent backcrosses (BC between hybrid males and parental females. This kind of sterility indueed through interspecific hybridization seems to be due to lack of sperm migration from the bursa copulatrix to the spermateca, thus suggesting primarily failure on the part of hybrid males to produce and/or to incorporate male accessory secretions into the spermatophore bulb. Addicional proof that sterility induced in hybrid males is at the sperm level has been afforded by the spermatogenesis herein studied. The anomalous processes like; 1 prophases of spermatogonia with the chromosomes scattered in the cytoplasm, 2 first metaphases with unpaired tetrades, 3 spermatids differing in size and 4 spermatozoa of abnormal shape and generdlly of giant size, can be taken as an indicator of the degree of departure from the normal course of spermatogenesis.

  20. 'Rogue' cells observed in children exposed to radiation from the Chernobyl accident

    International Nuclear Information System (INIS)

    Sevan'kaev, A.V.; Tsyb, A.F.; Zhloba, A.A.; Moiseenko, V.V.; Skrjabin, A.M.; Climov, V.M.

    1993-01-01

    Eight 'rogue' lymphocyte metaphases containing a large number of aberrant chromosomes were noted during a survey of chromosomal damage in 328 Belarussian children. The study population comprised children of families living in territory contaminated by radiation from the Chernobyl accident. The majority of the sample had been evacuated within 1 week from very heavily polluted territory to areas that had received much less fallout. Two hundred cells were scored per subject and one rogue cell was found in a child exposed in utero; one in a child conceived after the accident and six in the postnatally exposed group. The possibility that the damage was due to exposure to radio-iodine concentrated in the thyroid gland, or to radiation from incorporated hot particles' of an alpha or beta/gamma emitter is discussed. It is concluded that the damage to these cells is unlikely to have been caused by radiation. (Author)

  1. Nonrobertsonian translocation t(6;11) is associated with infertility in an oligoazoospermic male.

    Science.gov (United States)

    Pernice, Francesco; Mazza, Giuseppa; Puglisi, Domenico; Luppino, Maria Giuseppa; Frisina, Nicola

    2002-07-01

    To analyze an unusual nonrobertsonian translocation t(6;11) found in an infertile, oligoazoospermic, but otherwise apparently healthy man. Case report with review of the scientific literature. Academic research environment. Infertile, oligoazoospermic, but otherwise apparently healthy man. Peripheral blood lymphocytes were obtained for karyotyping, and metaphases were studied by standard GBG, RBG, and CBG banding procedures. Sperm count, morphology, and GBG, RBG, and CBG banding. A karyogram revealed a nonrobertsonian balanced translocation t(6;11) with breakpoints at 6q15 and 11p15, as shown by R-, G-, and C-banding. Physical contact of unpaired autosomal material with sex chromosomes, which leads to spermatogenic arrest, may be one of the factors that accounts for infertility in men with autosomal aberrations.

  2. Reply to Popescu and Zionjic

    Energy Technology Data Exchange (ETDEWEB)

    Jesudasan, R.A. [Center for Cellular and Molecular Biology, Hydrabad (India); Rahman, M.R. [Specialty Diagnosis, La Mirada, CA (United States); Evans, G.A. [Southwestern Medical School, Dallas, TX (United States); Srivatsan, E.S. [UCLA School of Medicine/VA Medical Center, West Los Angeles, CA (United States)

    1996-02-01

    In our studies, we analyzed eight different cervical cell lines, including two human papilloma virus (HPV)-negative cell lines, for rearrangements on chromosome 11. A number of approaches including RFLP, pulsed-field gel electrophoresis (PFGE), karyotyping, and FISH with 11 painting and 11q13-specific probes were used. Rearrangement of 11q13 sequences, translocation in two (HeLa and CasKi), deletion in one (SiHa), and translocation and deletion in one (C33A) were observed. One other cell line (HT3) showed rearrangement in PFGE gels. Metaphase spreads were not available for FISH studies on this cell line. Thus, 11q13 rearrangement was a nonrandom event. 8 refs.

  3. Live imaging of spindle pole disorganization in docetaxel-treated multicolor cells

    International Nuclear Information System (INIS)

    Sakaushi, Shinji; Nishida, Kumi; Minamikawa, Harumi; Fukada, Takashi; Oka, Shigenori; Sugimoto, Kenji

    2007-01-01

    Treatment of cells with docetaxel at low concentrations induces aberrant bipolar spindles of which two centrosomes stay at only one pole, and also induces multipolar spindles. To gain insight into the relations between centrosome impairment and structural defects of the spindle, live-cell imaging was performed on a human MDA Auro/imp/H3 cell line in which centrosomes/mitotic spindles, nuclear membrane and chromatin were simultaneously visualized by fluorescent proteins. In the presence of docetaxel at IC 50 concentration, the centrosomes did not segregate, and multiple aster-like structures ectopically arose around the disappearing nuclear membrane. Those ectopic structures formed an acentrosomal pole opposing to the two-centrosomes-containing pole. In late metaphase, one pole often fragmented into multiple spindle poles, leading multipolar division. These results suggest that spindle pole fragility may be induced by centrosome impairment, and collapse of the pole may contribute to induction of aneuploid daughter cells

  4. Euro-FISH: A concerted action towards standardized translocation detection in pathology

    DEFF Research Database (Denmark)

    Rijk, A. Van; Mason, D.; Leoncini, L.

    2007-01-01

    at a validation of a robust, standardized FISH protocol. When implemented this procedure will be an aid in discriminating the most difficult to diagnose cases. Method: The Euro-FISH protocol is a useful tool in discerning the different types of lymphoma and available treatment options. The Euro-FISH project......, organized in three stages therefore has established a robust standardized FISH protocol, to be used in routine laboratories for diagnostically difficult lymphoma cases. For the evaluation of the protocol 16 split-signal FISH probes were tested on 10 different lymphoma categories (MCL, splenic MZL, gastric...... MALT, CLL/SLL, DLBCL, FCL, LPL, BL, ALCL, T-LBL) supplemented with reactive tissues. The chromosomal localization of every probe was confirmed by using metaphase slides of healthy donors, before the probes were tested on the selected entities. Results and conclusions: Based on a shared protocol FISH...

  5. STATE-OF-THE-ART PRODUCTION OF EMBRYOS IN VITRO IN SOUTH AMERICAN CAMELIDS

    Directory of Open Access Journals (Sweden)

    Ruiz JB

    2015-12-01

    Full Text Available The first research on in vitro fertilization in camelids have been made using gametes of slaughtered animals recovered. This job has been of invaluable help to try to standardize protocols in vitro maturation and fertilization. However it has not yet determined the time of in vitro maturation of oocytes alpacas and llamas, recent research suggests that will require about 40 hours to obtain high rates of metaphase II oocytes. There have been several studies of in vitro fertilization of oocytes alpacas and llamas, there is still no defined protocol that will generate high rates of cleavage and blastocyst, however, pregnancies have been reported in llamas and alpacas and the birth of first breeding llama through IVF.

  6. Cytological investigation of Haplopappus gracilis (Nutt.) Gray: 5-methylcytosine-rich regions, fluorochrome banding and chromatin sensitivity to DNase I digestion.

    Science.gov (United States)

    Ruffini Castiglione, M; Frediani, M; Venora, G; Cremonini, R

    2008-01-01

    Haplopappus gracilis (Nutt.) Gray, one of the five known higher plants with a chromosome number of 2n = 4, was studied from a cytological point of view. The chromosome complement of this species was characterized by means of automated karyotype analysis. Moreover, the DNA methylation pattern and fluorochrome banding were determined and compared with cytological data present in the literature. DNA methylation distribution along metaphase chromosomes involved all chromosome territories evidenced by C-banding. Other methylated bands correlated positively with aceto-orcein-positive heterochromatic portions and/or with late replicating bands and/or fluorochrome bands. Some methylated bands showed differences between homologous chromosomes. These bands belonged partly to certain heterochromatic domains and partly to intercalary sites not defined by other standard banding techniques. Differences between the homologues were also indicated by our DNA content data obtained after DNase I digestion.

  7. C-heterochromatin and extra (B) chromosome distribution in six species of the Nabis (Heteroptera, Nabidae) with the modal male karyotype 2n = 16 + XY.

    Science.gov (United States)

    Grozeva, Snejana; Nokkala, Seppo

    2003-01-01

    The basic male karyotype of the six Nabis species (Heteroptera, Nabidae) is confirmed as being 2n=16+XY. The chromosomes are holokinetic while male meiosis is achiasmatic. The sex chromosomes undergo postreduction and in second metaphase show distance pairing, registered in all nabid species examined so far. Using C-banding technique for the first time in the family Nabidae, the heterochromatin was revealed on chromosomes of six species. The species showed different amount and distribution of C-heterochromatin. Only in Nabis (Dolichonabis) limbatus did the C-bands distribution make possible the identification of every chromosome pair in the karyotype. In other species, C-bands were found in some of the autosomes and the X, localized either interstitially or at telomeres. Only the Y usually showed relative stability ofthe C-banding pattern. In four of six species, extra (B) chromosomes were observed and their behaviour in meiosis described.

  8. SIRT1, 2, 3 protect mouse oocytes from postovulatory aging.

    Science.gov (United States)

    Zhang, Teng; Zhou, Yang; Li, Li; Wang, Hong-Hui; Ma, Xue-Shan; Qian, Wei-Ping; Shen, Wei; Schatten, Heide; Sun, Qing-Yuan

    2016-04-01

    The quality of metaphase II oocytes will undergo a time-dependent deterioration following ovulation as the result of the oocyte aging process. In this study, we determined that the expression of sirtuin family members (SIRT1, 2, 3) was dramatically reduced in mouse oocytes aged in vivo or in vitro. Increased intracellular ROS was observed when SIRT1, 2, 3 activity was inhibited. Increased frequency of spindle defects and disturbed distribution of mitochondria were also observed in MII oocytes aged in vitro after treatment with Nicotinamide (NAM), indicating that inhibition of SIRT1, 2, 3 may accelerate postovulatory oocyte aging. Interestingly, when MII oocytes were exposed to caffeine, the decline of SIRT1, 2, 3 mRNA levels was delayed and the aging-associated defective phenotypes could be improved. The results suggest that the SIRT1, 2, 3 pathway may play a potential protective role against postovulatory oocyte aging by controlling ROS generation.

  9. Genotoxic effect of formocresol pulp therapy of deciduous teeth.

    Science.gov (United States)

    Lucas Leite, Ana Catarina Gaioso; Rosenblatt, Aronita; da Silva Calixto, Merilane; da Silva, Cirlene Maria; Santos, Neide

    2012-08-30

    To investigate whether formocresol, in Buckley's original formulation, used for pulp therapy of deciduous teeth, can have a genotoxic effect. Genotoxicity was tested in lymphocyte cultures from the peripheral blood of children aged 5-10y, in Recife, Pernambuco, Brazil. This was a case-control study. The sample comprised 40 children who had primary teeth with non-vital pulps. Two venous blood samples (6-8ml) were collected from each child, the first prior to pulp therapy (control group) and the second 24h after pulp therapy (experimental group). Lymphocyte cultures were grown in 78% RPMI 1640 medium, 20% fetal bovine serum, 2% phytohemagglutinin. The lymphocytes were assessed for chromosomal aberrations; each sample involved analysis of 100 metaphases. There was a statistically significant difference between the control and treated groups for the isochromatid gap (pformocresol in pediatric dentistry is recommended. Copyright © 2012 Elsevier B.V. All rights reserved.

  10. Evaluation of the genotoxic effects of formocresol application in vital pulp therapy of primary teeth: a clinical study and meta-analysis.

    Science.gov (United States)

    de Alencar Filho, Arnoldo Vasconcelos; Dos Santos Junior, Valdeci Elias; da Silva Calixto, Merilane; Santos, Neide; Heimer, Monica Vilela; Rosenblatt, Aronita

    2018-02-06

    This in vivo research investigated whether pulp treatments using formocresol for 7 days would cause mutagenic changes in children's lymphocytes. The mutagenicity was tested in lymphocyte cultures established from the peripheral blood of children living in Brazil. The samples consisted of 2000 cells from teeth undergoing formocresol pulpotomies in which the formocresol pellet was sealed in the primary tooth for 7 days. It was removed on the seventh day, the base was placed, and the tooth was restored. Two venous blood samples (6-8 ml) were collected from each child; the first was prior to pulp therapy, and the second was 7 days later. Two thousand metaphases were analyzed. The level of significance adopted for the statistics was P formocresol. This research helps to bring scientific evidence of the safe use of formocresol in deciduous pulpotomy treatments.

  11. Unusual arrangement and behaviour of the sex chromosomes of Aphodius (Agolius abdominalis Bonelli, 1812, and comparison with A. (A. bonvouloiri Harold, 1860 (Coleoptera: Aphodiidae

    Directory of Open Access Journals (Sweden)

    Robert Angus

    2009-12-01

    Full Text Available Aphodius abdominalis Bonelli, 1812 is shown to have a karyotype comprising nine pairs of autosomes and sex chromosomes which are X0 (male, XX (female. At first metaphase of meiosis the X chromosome is linked to an autosomal bivalent by a darkly staining area of the cytoplasm, resembling the Xy p arrangement typical of Aphodius species, but giving nine, rather than 10, elements in the nucleus. C-banding, which shows the centromeres, confirms this unusual arrangement. A. bonvouloiri, the only other known species of subgenus Agolius Mulsant et Rey, 1869, has a male karyotype with nine pairs of autosomes and Xy sex chromosomes. No preparations of its meiosis are available.

  12. Cyclin A2 modulates kinetochore–microtubule attachment in meiosis II

    Science.gov (United States)

    Yuen, Wai Shan; Adhikari, Deepak; FitzHarris, Greg; Conti, Marco; Sicinski, Piotr; Nabti, Ibtissem; Marangos, Petros

    2017-01-01

    Cyclin A2 is a crucial mitotic Cdk regulatory partner that coordinates entry into mitosis and is then destroyed in prometaphase within minutes of nuclear envelope breakdown. The role of cyclin A2 in female meiosis and its dynamics during the transition from meiosis I (MI) to meiosis II (MII) remain unclear. We found that cyclin A2 decreases in prometaphase I but recovers after the first meiotic division and persists, uniquely for metaphase, in MII-arrested oocytes. Conditional deletion of cyclin A2 from mouse oocytes has no discernible effect on MI but leads to disrupted MII spindles and increased merotelic attachments. On stimulation of exit from MII, there is a dramatic increase in lagging chromosomes and an inhibition of cytokinesis. These defects are associated with an increase in microtubule stability in MII spindles, suggesting that cyclin A2 mediates the fidelity of MII by maintaining microtubule dynamics during the rapid formation of the MII spindle. PMID:28819014

  13. Genome Transfer Prevents Fragmentation and Restores Developmental Potential of Developmentally Compromised Postovulatory Aged Mouse Oocytes

    Directory of Open Access Journals (Sweden)

    Mitsutoshi Yamada

    2017-03-01

    Full Text Available Changes in oocyte quality can have great impact on the developmental potential of early embryos. Here we test whether nuclear genome transfer from a developmentally incompetent to a developmentally competent oocyte can restore developmental potential. Using in vitro oocyte aging as a model system we performed nuclear transfer in mouse oocytes at metaphase II or at the first interphase, and observed that development to the blastocyst stage and to term was as efficient as in control embryos. The increased developmental potential is explained primarily by correction of abnormal cytokinesis at anaphase of meiosis and mitosis, by a reduction in chromosome segregation errors, and by normalization of the localization of chromosome passenger complex components survivin and cyclin B1. These observations demonstrate that developmental decline is primarily due to abnormal function of cytoplasmic factors involved in cytokinesis, while the genome remains developmentally fully competent.

  14. From Meiosis to Mitosis: The Astonishing Flexibility of Cell Division Mechanisms in Early Mammalian Development.

    Science.gov (United States)

    Bury, L; Coelho, P A; Glover, D M

    2016-01-01

    The execution of female meiosis and the establishment of the zygote is arguably the most critical stage of mammalian development. The egg can be arrested in the prophase of meiosis I for decades, and when it is activated, the spindle is assembled de novo. This spindle must function with the highest of fidelity and yet its assembly is unusually achieved in the absence of conventional centrosomes and with minimal influence of chromatin. Moreover, its dramatic asymmetric positioning is achieved through remarkable properties of the actin cytoskeleton to ensure elimination of the polar bodies. The second meiotic arrest marks a uniquely prolonged metaphase eventually interrupted by egg activation at fertilization to complete meiosis and mark a period of preparation of the male and female pronuclear genomes not only for their entry into the mitotic cleavage divisions but also for the imminent prospect of their zygotic expression. © 2016 Elsevier Inc. All rights reserved.

  15. Epidemiological study using the chromosome aberration technique in different population samples

    International Nuclear Information System (INIS)

    Jesus Prieto, M.; Moreno, M.; Olivares, P.; Gomez, M.; Herranz, R.

    1997-01-01

    As far back as 10 years ago, the Gregorio Maranon General University Hospital (HGUGM) was equipped with a laboratory of biological dosimetry and has undertaken dosimetric estimations of individuals suspected with overexposure to ionizing radiation. For this purpose it was necessary to carry out studies on the basic frequency of chromosomal aberrations in a controlled (radiation exposed) population. A study was conducted on 72 individuals of the community of Madrid who had not been exposed to ionizing radiation. 500 metaphases per individual were examined and a basic frequency of 0.7 (dic)/1000 cells analysed. Parameters such as age, gender and cigarette consumption were considered and results show a linear dependency with age and consumption of cigarettes, but not with gender

  16. Effect of beta-mercaptoethanol or epidermal growth factor supplementation on in vitro maturation of canine oocytes collected from dogs with different stages of the estrus cycle.

    Science.gov (United States)

    Kim, Min Kyu; Fibrianto, Yuda Heru; Oh, Hyun Ju; Jang, Goo; Kim, Hye Jin; Lee, Kyu Seung; Kang, Sung Keun; Lee, Byeong Chun; Hwang, Woo Suk

    2004-09-01

    Supplementation of beta-mercaptoethanol (beta-ME) in in vitro maturation (IVM) medium was shown to improve embryo development and quality in several species. Epidermal growth factor (EGF) was also shown to improve IVM of human oocyte and embryo development after in vitro fertilization (IVF). The effect of these two compounds were suggested to be mediated through the synthesis of glutathione (GSH) which is known to play an important role in protecting the cell or embryos from oxidative damage. Thus, it is suggested that supplementation of canine IVM medium with beta-ME or EGF may be of benefit due to its positive role in IVM of various mammalian oocytes and embryo development, including cattle, pigs, rodents and humans. This study investigates the effect of ovarian estrus stage on canine oocyte quality and supplementation of medium with beta-ME or EGF on IVM of canine oocytes. As results, a significantly higher percentage of oocytes progressed to metaphase II (MII) stage in 50 or 100 microM of beta-ME supplemented oocytes collected from the follicular stage. The maturation rate to metaphase I (MI) stage was also significantly higher in oocytes collected from follicular stage and cultured with 25 or 100 microM compared to other experimental groups. After IVM culture, oocytes recovered from dogs with the follicular stage and matured in TCM-199 supplemented with 20 ng/ml EGF yielded better oocyte maturation to MII phase compared to other groups. Taken together, supplementation of beta-ME (50 or 100 microM) or EGF (20 ng/ml) improved IVM of canine oocytes to MII stage.

  17. Separase loss of function cooperates with the loss of p53 in the initiation and progression of T- and B-cell lymphoma, leukemia and aneuploidy in mice.

    Directory of Open Access Journals (Sweden)

    Malini Mukherjee

    Full Text Available Cohesin protease Separase plays a key role in faithful segregation of sister chromatids by cleaving the cohesin complex at the metaphase to anaphase transition. Homozygous deletion of ESPL1 gene that encodes Separase protein results in embryonic lethality in mice and Separase overexpression lead to aneuploidy and tumorigenesis. However, the effect of Separase haploinsufficiency has not been thoroughly investigated.Here we examined the effect of ESPL1 heterozygosity using a hypomorphic mouse model that has reduced germline Separase activity. We report that while ESPL1 mutant (ESPL1 (+/hyp mice have a normal phenotype, in the absence of p53, these mice develop spontaneous T- and B-cell lymphomas, and leukemia with a significantly shortened latency as compared to p53 null mice. The ESPL1 hypomorphic, p53 heterozygous transgenic mice (ESPL1(+/hyp, p53(+/- also show a significantly reduced life span with an altered tumor spectrum of carcinomas and sarcomas compared to p53(+/- mice alone. Furthermore, ESPL1(+/hyp, p53(-/- mice display significantly higher levels of genetic instability and aneuploidy in normal cells, as indicated by the abnormal metaphase counts and SKY analysis of primary splenocytes.Our results indicate that reduced levels of Separase act synergistically with loss of p53 in the initiation and progression of B- and T- cell lymphomas, which is aided by increased chromosomal missegregation and accumulation of genomic instability. ESPL1(+/hyp, p53(-/- mice provide a new animal model for mechanistic study of aggressive lymphoma and also for preclinical evaluation of new agents for its therapy.

  18. Enhancing Cytogenetic Biological Dosimetry Capabilities of the Philippines for Nuclear Incident Preparedness.

    Science.gov (United States)

    Asaad, Celia O; Caraos, Gloriamaris L; Robles, Gerardo Jose M; Asa, Anie Day D C; Cobar, Maria Lucia C; Asaad, Al-Ahmadgaid

    2016-01-01

    The utility of a biological dosimeter based on the analysis of dicentrics is invaluable in the event of a radiological emergency wherein the estimated absorbed dose of an exposed individual is crucial in the proper medical management of patients. The technique is also used for routine monitoring of occupationally exposed workers to determine radiation exposure. An in vitro irradiation study of human peripheral blood lymphocytes was conducted to establish a dose-response curve for radiation-induced dicentric aberrations. Blood samples were collected from volunteer donors and together with optically stimulated luminescence (OSL) dosimeters and were irradiated at 0, 0.1, 0.25, 0.5, 0.75, 1, 2, 4, and 6 Gy using a cobalt-60 radiotherapy unit. Blood samples were cultured for 48 h, and the metaphase chromosomes were prepared following the procedure of the International Atomic Energy Agency's Emergency Preparedness and Response - Biodosimetry 2011 manual. At least 100 metaphases were scored for dicentric aberrations at each dose point. The data were analyzed using R language program. The results indicated that the distribution of dicentric cells followed a Poisson distribution and the dose-response curve was established using the estimated model, Y dic = 0.0003 (±0.0003) +0.0336 (±0.0115) × D + 0.0236 (±0.0054) × D 2 . In this study, the reliability of the dose-response curve in estimating the absorbed dose was also validated for 2 and 4 Gy using OSL dosimeters. The data were fitted into the constructed curve. The result of the validation study showed that the obtained estimate for the absorbed exposure doses was close to the true exposure doses.

  19. A high incidence of meiotic silencing of unsynapsed chromatin is not associated with substantial pachytene loss in heterozygous male mice carrying multiple simple robertsonian translocations.

    Directory of Open Access Journals (Sweden)

    Marcia Manterola

    2009-08-01

    Full Text Available Meiosis is a complex type of cell division that involves homologous chromosome pairing, synapsis, recombination, and segregation. When any of these processes is altered, cellular checkpoints arrest meiosis progression and induce cell elimination. Meiotic impairment is particularly frequent in organisms bearing chromosomal translocations. When chromosomal translocations appear in heterozygosis, the chromosomes involved may not correctly complete synapsis, recombination, and/or segregation, thus promoting the activation of checkpoints that lead to the death of the meiocytes. In mammals and other organisms, the unsynapsed chromosomal regions are subject to a process called meiotic silencing of unsynapsed chromatin (MSUC. Different degrees of asynapsis could contribute to disturb the normal loading of MSUC proteins, interfering with autosome and sex chromosome gene expression and triggering a massive pachytene cell death. We report that in mice that are heterozygous for eight multiple simple Robertsonian translocations, most pachytene spermatocytes bear trivalents with unsynapsed regions that incorporate, in a stage-dependent manner, proteins involved in MSUC (e.g., gammaH2AX, ATR, ubiquitinated-H2A, SUMO-1, and XMR. These spermatocytes have a correct MSUC response and are not eliminated during pachytene and most of them proceed into diplotene. However, we found a high incidence of apoptotic spermatocytes at the metaphase stage. These results suggest that in Robertsonian heterozygous mice synapsis defects on most pachytene cells do not trigger a prophase-I checkpoint. Instead, meiotic impairment seems to mainly rely on the action of a checkpoint acting at the metaphase stage. We propose that a low stringency of the pachytene checkpoint could help to increase the chances that spermatocytes with synaptic defects will complete meiotic divisions and differentiate into viable gametes. This scenario, despite a reduction of fertility, allows the spreading

  20. The Latin American Biological Dosimetry Network (LBDNet): Argentina, Brazil, Chile, Cuba, Mexico, Peru, Uruguay

    International Nuclear Information System (INIS)

    Guerrero C, C.; Arceo M, C.; Di Giorgio, M.; Vallerga, M.; Radl, A.; Taja, M.; Seoane, A.; De Luca, J.; Stuck O, M.; Valdivia, P.

    2010-10-01

    Biological dosimetry is a necessary support for national radiation protection programs and emergency response schemes. The Latin American Biological Dosimetry Network (LBDNet) was formally founded in 2007 for mutual assistance in case of radiation emergencies and for providing support to other Latin American countries that do not have bio dosimetry laboratories. In the frame of the IAEA Technical Cooperation Projects RLA/9/54 and RLA/9/61 the following activities have been performed: a) An international intercomparison exercise organized during 2007-2008 included six European countries and LBDNet laboratories. Relevant parameters related with dose assessment were evaluated through triage and conventional scoring criteria. A new approach for statistical data analysis was developed including assessment of inter-laboratory reproducibility and intra-laboratory repeatability. Overall, the laboratory performance was satisfactory for mutual cooperation purposes. b) In 2009, LBDNet and two European countries carried out a digital image intercomparison exercise involving dose assessment from metaphase images distributed electronically through internet. The main objectives were to evaluate scoring feasibility on metaphase images and time response. In addition a re-examination phase was considered in which the most controversial images were discussed jointly, this allowed for the development of a homogeneous scoring criteria within the network. c) A further exercise was performed during 2009 involving the shipment of biological samples for biological dosimetry assessment. The aim of this exercise was to test the timely and properly sending and receiving blood samples under national and international regulations. A total of 14 laboratories participated in this joint IAEA, PAHO and WHO. (Author)

  1. Diabetes Insipidus as an Initial Presentation of Myelodysplastic Syndrome: Diagnosis with Single-Nucleotide Polymorphism Array-Based Karyotyping.

    Science.gov (United States)

    Sun, Ruixue; Wang, Chun; Zhong, Xushu; Wu, Yu

    2016-04-01

    Myelodysplastic syndrome (MDS) is a group of clonal hematopoietic diseases characterized by cytopenia, dysplasia and increased risk of development to acute myeloid leukemia (AML). Unfavorable cytogenetic changes such as complex karyotypes or chromosome 7 anomalies are predictive of the progression to AML and poor prognosis. Central diabetes insipidus (CDI) is the result of a deficiency of arginine vasopressin, and its major causes are idiopathic, primary or secondary tumors, neurosurgery and trauma. Importantly, CDI is a rare complication of MDS. To date, only 5 cases of MDS co-occurring with CDI have been reported; 3 of 5 had cytogenetic abnormalities uncovered by metaphase cytogenetics and 3 of 5 evolved to AML. Here, we describe a 74-year-old woman who presented with CDI as her initial symptom of MDS and eventually progressed to AML. The metaphase cytogenetics, combined with the single-nucleotide polymorphism array (SNP-A)-based karyotyping, with superiority in resolution and detecting copy number variation, revealed a complex karyotype that included monosomy of chromosome 7, deletion of 20q, and absence of heterogeneity (AOH) in more than one chromosome. To the best of our knowledge, this is the first case report of MDS co-occurring with CDI with numerous cytogenetic abnormalities revealed by the SNP-A-based karyotyping. Our case supports that the cytogenetic abnormalities may be associated with the clinical features and the prognosis of MDS co-occurring with CDI. The SNP-A-based karyotyping is helpful in revealing more subtle cytogenetic abnormalities and unveiling their roles in the pathogenesis of MDS.

  2. In vitro maturation and embryo development of bovine oocytes after meiosis blockage with MPF inhibitors

    Directory of Open Access Journals (Sweden)

    Mariana Groke Marques

    2011-12-01

    Full Text Available This study evaluated the bovine oocyte maturation and embryo development after in vitro fertilization. The maturation of the oocytes was blocked using Butyrolactone I and Roscovitine using pre-maturation medium supplemented with fetal calf serum (FCS. The ocytes were divided in four groups: Control 0 hour, Control (24 hours of maturation, Roscovitine (maturation blockage with 50mM Roscovitine during 24 hours followed by 24 hours of maturation, and Butyrolactone I (maturation blockage with 150mM Butyrolactone I during 24 hours followed by 24 hours of maturation. The oocytes were fixed and stained with aceto orcein to evaluate the nuclear maturation. After the maturation period, the remaining oocytes of the Control group, Roscovitine, and Butyrolactone I were fertilized in vitro. Embryo development was assessed by the cleavage rate (D3 and blastocysts formation (D7. The Butyrolactone I group had similar rates of germinal vesical stage oocytes during blockage, and Metaphase 2 after maturation, comparing to Control group at 0 hour and Control group, respectively. On the other hand, the Roscovitine group had lower rates of vesical stage oocytes during blockage, and Metaphase 2 after maturation comparing to Control groups. After in vitro fertilization, higher rates of cleavage were observed in Control and Butyrolactone I groups. For the blastocyst formation rate, the Control group showed better results than Roscovitine group. In summary, Butyrolactone I group had similar results to the Control group, and for this reason, is suitable for pre-maturation of bovine oocytes using FCS. In contrast, Roscovitine group had lower oocyte maturation and embryo development.

  3. Karyotype Learning Center: A Software For Teaching And Learning Cytogenetics

    Directory of Open Access Journals (Sweden)

    Joelma Freire De Mesquita

    2004-05-01

    Full Text Available The in vitro cultivation of human cells is an essential part of the work of every diagnostic cytoge-netics laboratory. Almost all human cytogenetic studies involve the examination of dividing bloodcell population by blocking cell division at metaphase with subsequent processing and staining bybanding techniques. The chromosome constitution is described as Karyotype that states the totalnumber of chromosomes and the sex chromosome constitution. Karyotypes are prepared by cuttingup a photograph of the spread metaphase chromosomes, matching up homologous chromosomes andsticking them back down on a card or nowadays more often by getting an image analysis computerto do the job. Chromosomes are identied by their size, centromere position and banding pattern.Teaching a student how to detect and interpret even the most common chromosome abnormaliti-es is a major challenge: mainly, in a developing country where the laboratorial facilities are notalways available for a big number of students. Therefore, in this work we present an educationalsoftware for teaching undergraduate students of Medical and Life Sciences Courses how to arrangenormal and abnormal chromosomes in the form of karyotype. The user, using drag-and-drop, is da-red to match up homologous chromosome. For that, we have developed a free full access web site(http://www.biomol.net/cariotipo/ for hosting the software. The latter has proved to be light andfast even under slow dial-up connections. This web site also oers a theoretical introductory sectionwith basic concepts about karyotype. Up to now the software has been successfully applied to un-dergraduate courses at the University of Rio de Janeiro (UNIRIO. The students have approved thesoftware; to them the similarities with the well-known game solitaire turns the exercise more excitingand provides additional stimulus to learn and understand karyotype. Professors have also used thesoftware as complementary material in their regular classes

  4. Radiolabeled blood cells: radiation dosimetry and significance

    International Nuclear Information System (INIS)

    Thakur, M.L.

    1986-01-01

    Over the past few years blood cells labeled with In-111 have become increasingly useful in clinical diagnosis and biomedical research. Indium-111 by the virtue of its physical characteristics and ability to bind to cell cytoplasmic components, provides an excellent cell tracer and thereby, allows investigators to monitor in vivo cell distribution by external imaging and help determine a course of regimen in treating life threatening diseases. Due to natural phenomena such as margination, blood pool, and reticuloendothelial cell activity, in the normal state, depending upon the cell type and the quality of cell preparations, 30%-50% of the administered radioactivity is immediately distributed in the liver, spleen and bone marrow. Over a period of time the radioactivity in these organs slightly increases and decays with a physical half-life of In-111. The resulting radiation dose to these organs ranges between 1-25 rads/mCi In-111 administered. The authors have developed a new In-111 labeling technique which preserves platelet ultrastructure and shown that human lymphocytes labeled with In-111 in mixed leukocytes preparations a) are only 0.003% of the total -body lymphocytes population and b) are killed. The consequence if any may be considered insignificant, particularly because 5.6% metaphases from normal men and 6.5% metaphases from normal women in the US have at least one chromosome aberration. Calculations have shown that the risk of fatal hematological malignancy, over a 30 year period, in recipients of 100 million lymphocytes labeled with 100 μCi In-111 is 1/million patients studied. This risk is less than 0.025% of the 1981 spontaneous cancer patient rate in the country. 32 references, 10 tables

  5. Basal aurora kinase B activity is sufficient for histone H3 phosphorylation in prophase

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    Ly-Thuy-Tram Le

    2013-02-01

    Histone H3 phosphorylation is the hallmark of mitosis deposited by aurora kinase B. Benzo[e]pyridoindoles are a family of potent, broad, ATP-competitive aurora kinase inhibitors. However, benzo[e]pyridoindole C4 only inhibits histone H3 phosphorylation in prophase but not in metaphase. Under the C4 treatment, the cells enter into mitosis with dephosphorylated histone H3, assemble chromosomes normally and progress to metaphase, and then to anaphase. C4 also induces lagging chromosome in anaphase but we demonstrated that these chromosome compaction defects are not related to the absence of H3 phosphorylation in prophase. As a result of C4 action, mitosis lasts longer and the cell cycle is slowed down. We reproduced the mitotic defects with reduced concentrations of potent pan aurora kinase as well as with a specific aurora B ATP-competitive inhibitor; we therefore propose that histone H3 phosphorylation and anaphase chromosome compaction involve the basal activity of aurora kinase B. Our data suggest that aurora kinase B is progressively activated at mitosis entry and at anaphase onset. The full activation of aurora kinase B by its partners, in prometaphase, induces a shift in the catalytic domain of aurora B that modifies its affinity for ATP. These waves of activation/deactivation of aurora B correspond to different conformations of the chromosomal complex revealed by FRAP. The presence of lagging chromosomes may have deleterious consequences on the daughter cells and, unfortunately, the situation may be encountered in patients receiving treatment with aurora kinase inhibitors.

  6. Smurf2 as a novel mitotic regulator: From the spindle assembly checkpoint to tumorigenesis

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    Moore Finola E

    2009-07-01

    Full Text Available Abstract The execution of the mitotic program with high fidelity is dependent upon precise spatiotemporal regulation of posttranslational protein modifications. For example, the timely polyubiquitination of critical mitotic regulators by Anaphase Promoting Complex/Cyclosome (APC/C is essential for the metaphase to anaphase transition and mitotic exit. The spindle assembly checkpoint prevents unscheduled activity of APC/C-Cdc20 in early mitosis, allowing bipolar attachment of kinetochores to mitotic spindle and facilitating equal segregation of sister chromatids. The critical effector of the spindle checkpoint, Mitotic arrest deficient 2 (Mad2, is recruited to unattached kinetochores forming a complex with other regulatory proteins to efficiently and cooperatively inhibit APC/C-Cdc20. A weakened and/or dysfunctional spindle checkpoint has been linked to the development of genomic instability in both cell culture and animal models, and evidence suggests that aberrant regulation of the spindle checkpoint plays a critical role in human carcinogenesis. Recent studies have illuminated a network of both degradative and non-degradative ubiquitination events that regulate the metaphase to anaphase transition and mitotic exit. Within this context, our recent work showed that the HECT (Homologous to E6-AP C-terminus-family E3 ligase Smurf2 (Smad specific ubiquitin regulatory factor 2, known as a negative regulator of transforming growth factor-beta (TGF-β signaling, is required for a functional spindle checkpoint by promoting the functional localization and stability of Mad2. Here we discuss putative models explaining the role of Smurf2 as a new regulator in the spindle checkpoint. The dynamic mitotic localization of Smurf2 to the centrosome and other critical mitotic structures provides implications about mitotic checkpoint control dependent on various ubiquitination events. Finally, deregulated Smurf2 activity may contribute to carcinogenesis by

  7. Hypermutability of mouse chromosome 2 during the development of x-ray-induced murine myeloid leukemia

    International Nuclear Information System (INIS)

    Rithidech, K.; Bond, V.P.; Cronkite, E.P.; Thompson, M.H.; Bullis, J.E.

    1995-01-01

    In an effort to identify the precise role of a deletion at regions D-E of mouse chromosome 2 [del2(D-E)] during the development of radiation-induced myeloid leukemia, we conducted a serial sacrifice study in which metaphase chromosomes were examined by the G-banding technique. Such metaphase cells were collected from x-irradiated mice during the period of transformation of some of the normal hematopoietic cells to the fully developed leukemic phenotype. A group of 250 CBA/Ca male mice (10-12 weeks old) were exposed to a single dose of 2 Gy of 250-kilovolt-peak x-rays; 42 age-matched male mice served as controls. Groups of randomly selected mice were sacrificed at 20 hr, 1 week, and then at intervals of 3 months up to 24 months after x-irradiation. Slides for cytogenetic, hematological, and histological examination were prepared for each animal at each sacrifice time. The majority of such lesions were translocations at 2F or 2H, strongly suggesting hyper mutability of these sites on mouse chromosome 2. No lesions were found in control mice. The finding leads to the possibility that genomic lesions close to 2D and 2E are aberrants associated with radiation leukemogenesis, whereas a single clone of cells with a del2(D-E) may lead directly to overt leukemia. The data also indicate that leukemic transformation arises from the cumulative effects of multiple genetic events on chromosome 2, reinforcing the thesis that multiple steps of mutation occur in the pathogenesis of cancer. 15 refs., 1 fig., 2 tabs

  8. Telomeres and their possible role in chromosome stabilization

    Energy Technology Data Exchange (ETDEWEB)

    Day, J.P.; Marder, B.A.; Morgan, W.F. (Univ. of California, San Francisco, CA (United States))

    1993-01-01

    The evidence to date generally supports the hypothesis that telomere capping makes chromosome fragments refractory to subsequent rejoining events, but this control may be somewhat relaxed after chromosome breakage. Cell survival requires that the fragments rejoin before metaphase. Unprotected ends such as those produced by DNA damage are subject to degradation, presumably by endogenous cellular exo- and endonucleases. Telomere repeat sequences may be added to broken chromosome ends to protect the ends from further degradation. That telomeric DNA does not always prevent rejoining raises interesting questions as to what constitutes capping, and how rapidly it occurs after DNA damage in relation to chromosome break rejoining. The prevention of degradation and control of rejoining may be mediated by telomere-specific binding proteins, especially the telomere terminal binding protein. Some of these proteins may be involved in scavenging telomeric DNA when the cell senses that chromosomal breaks have occurred. Although chromosome break rejoining is an efficient process in eukaryotic cells, some breaks are never rejoined and can result in terminal delections and chromatid and isochromatid deletions at metaphase. It is unclear why these breaks are not rejoined, but it may be due to one or more of the following: (1) chance: broken chromosomes are separated, do not approach sufficiently close to one another, and are consequently physically unable to rejoin; (2) a large number of added telomere repeat sequences indicating to the cell that the chromosome has an authentic telomere; (3) some other DNA modification event that protects DNA ends from degradation, e.g., folding back of DNA ends to form a hairpin, as has been implicated in VDJ recombination.

  9. [Cytogenetic effect of cyclophosphamide in a culture of human lymphocytes following its activation in the bodies of mice].

    Science.gov (United States)

    Chebotarev, A N; Telegin, L I; Derzhavets, E M

    1976-01-01

    Cytogenetic effect of cyclophosphamide in cultured human lymphocytes after its activation in C57BL/6 mice in vivo was investigated. Cyclophosphamide was injected intraperitoneally in mice for 30 min. at doses of 200, 400, 600, 800 and 1000 mg/kg. Blood serum with activated metabolites of cyclophosphamide was added to human lymphocyte culture. The dependence of the part of aberrant metaphases on the concentration of cyclophosphamide after the activation can be presented as equation rho==1-e-(KC+alpha)2 and the total number of breaks as X=e(KC+alpha)2-1, where rho is a part of aberrant metaphases, X is a number of breaks of chromosomes per cell, C is the concentration, K and alpha are coefficients. The part of chromatid breaks from the total number of chromosome damages is constant for all concentrations and the comprises on the average 79,11%. Only the chromatid type of exchanges are observed. Distribution of chromosome breaks in cells corresponds to geometrical, but not to Poisson's distribution. Cyclophosphamide belongs to the group of one-sited mutagens in its cytogenetic chatacteristics. The alkylating activity of cyclophosphamide metabolites, estimated by means of NBP test, increases up to the dose 400 mg/kg and then remains constant for the strain of mice studied, cytogenetic activity increasing. Cyclophosphamide does not produce cytogenetic activity without activation. To test chemical substances for mutagenic activity, it is suggested to activate them in the mouse organism with the following administrating blood serum of these animals with the metabolites of tested (or with primary) substances in the study of their mutagenic activity on human lymphocyte culture.

  10. Cytoplasmic MTOCs control spindle orientation for asymmetric cell division in plants.

    Science.gov (United States)

    Kosetsu, Ken; Murata, Takashi; Yamada, Moé; Nishina, Momoko; Boruc, Joanna; Hasebe, Mitsuyasu; Van Damme, Daniël; Goshima, Gohta

    2017-10-17

    Proper orientation of the cell division axis is critical for asymmetric cell divisions that underpin cell differentiation. In animals, centrosomes are the dominant microtubule organizing centers (MTOC) and play a pivotal role in axis determination by orienting the mitotic spindle. In land plants that lack centrosomes, a critical role of a microtubular ring structure, the preprophase band (PPB), has been observed in this process; the PPB is required for orienting (before prophase) and guiding (in telophase) the mitotic apparatus. However, plants must possess additional mechanisms to control the division axis, as certain cell types or mutants do not form PPBs. Here, using live imaging of the gametophore of the moss Physcomitrella patens , we identified acentrosomal MTOCs, which we termed "gametosomes," appearing de novo and transiently in the prophase cytoplasm independent of PPB formation. We show that gametosomes are dispensable for spindle formation but required for metaphase spindle orientation. In some cells, gametosomes appeared reminiscent of the bipolar MT "polar cap" structure that forms transiently around the prophase nucleus in angiosperms. Specific disruption of the polar caps in tobacco cells misoriented the metaphase spindles and frequently altered the final division plane, indicating that they are functionally analogous to the gametosomes. These results suggest a broad use of transient MTOC structures as the spindle orientation machinery in plants, compensating for the evolutionary loss of centrosomes, to secure the initial orientation of the spindle in a spatial window that allows subsequent fine-tuning of the division plane axis by the guidance machinery. Copyright © 2017 the Author(s). Published by PNAS.

  11. Chromosomal Behavior during Meiosis in the Progeny of Triticum timopheevii × Hexaploid Wild Oat

    Science.gov (United States)

    An, Hongzhou; Hu, Mei; Li, Pengfei; Geng, Guangdong; Zhang, Qingqin; Zhang, Suqin

    2015-01-01

    The meiotic behavior of pollen mother cells (PMCs) of the F2 and F3 progeny from Triticum timopheevii × hexaploid wild oat was investigated by cytological analysis and sequential C-banding-genomic in situ hybridization (GISH) in the present study. A cytological analysis showed that the chromosome numbers of the F2 and F3 progeny ranged from 28 to 41. A large number of univalents, lagging chromosomes, chromosome bridges and micronuclei were found at the metaphase I, anaphase I, anaphase II and tetrad stages in the F2 and F3 progeny. The averages of univalents were 3.50 and 2.73 per cell, and those of lagging chromosomes were 3.37 and 1.87 in the F2 and F3 progeny, respectively. The PMC meiotic indices of the F2 and F3 progeny were 12.22 and 20.34, respectively, indicating considerable genetic instability. A sequential C-banding-GISH analysis revealed that some chromosomes and fragments from the hexaploid wild oat were detected at metaphase I and anaphase I in the progeny, showing that the progeny were of true intergeneric hybrid origin. The alien chromosomes 6A, 7A, 3C and 2D were lost during transmission from F2 to F3. In addition, partial T. timopheevii chromosomes appeared in the form of univalents or lagging chromosomes, which might result from large genome differences between the parents, and the wild oat chromosome introgression interfered with the wheat homologues’ normally pairing. PMID:25950431

  12. Chromosomal Behavior during Meiosis in the Progeny of Triticum timopheevii × Hexaploid Wild Oat.

    Directory of Open Access Journals (Sweden)

    Hongzhou An

    Full Text Available The meiotic behavior of pollen mother cells (PMCs of the F2 and F3 progeny from Triticum timopheevii × hexaploid wild oat was investigated by cytological analysis and sequential C-banding-genomic in situ hybridization (GISH in the present study. A cytological analysis showed that the chromosome numbers of the F2 and F3 progeny ranged from 28 to 41. A large number of univalents, lagging chromosomes, chromosome bridges and micronuclei were found at the metaphase I, anaphase I, anaphase II and tetrad stages in the F2 and F3 progeny. The averages of univalents were 3.50 and 2.73 per cell, and those of lagging chromosomes were 3.37 and 1.87 in the F2 and F3 progeny, respectively. The PMC meiotic indices of the F2 and F3 progeny were 12.22 and 20.34, respectively, indicating considerable genetic instability. A sequential C-banding-GISH analysis revealed that some chromosomes and fragments from the hexaploid wild oat were detected at metaphase I and anaphase I in the progeny, showing that the progeny were of true intergeneric hybrid origin. The alien chromosomes 6A, 7A, 3C and 2D were lost during transmission from F2 to F3. In addition, partial T. timopheevii chromosomes appeared in the form of univalents or lagging chromosomes, which might result from large genome differences between the parents, and the wild oat chromosome introgression interfered with the wheat homologues' normally pairing.

  13. The Latin American Biological Dosimetry Network (LBDNet): Argentina, Brazil, Chile, Cuba, Mexico, Peru, Uruguay

    Energy Technology Data Exchange (ETDEWEB)

    Guerrero C, C.; Arceo M, C. [ININ, Carretera Mexico-Toluca s/n, Ocoyoacac 52750, Estado de Mexico (Mexico); Di Giorgio, M.; Vallerga, M.; Radl, A. [Autoridad Regulatoria Nuclear, Av. del Libertador 8250, C1429 BNP CABA (Argentina); Taja, M.; Seoane, A.; De Luca, J. [Universidad Nacionald de La Plata, Av. 7 No. 1776, La Plata 1900, Buenos Aires (Argentina); Stuck O, M. [Instituto de Radioproteccion y Dosimetria, Av. Salvador Allende s/n, Recreio dos Bandeirantes, Rio de Janeiro (Brazil); Valdivia, P., E-mail: lbdnet@googlegroups.co [Comision Chilena de Energia, Amutanegui 95, Santiago Centro, Santiago (Chile)

    2010-10-15

    Biological dosimetry is a necessary support for national radiation protection programs and emergency response schemes. The Latin American Biological Dosimetry Network (LBDNet) was formally founded in 2007 for mutual assistance in case of radiation emergencies and for providing support to other Latin American countries that do not have bio dosimetry laboratories. In the frame of the IAEA Technical Cooperation Projects RLA/9/54 and RLA/9/61 the following activities have been performed: a) An international intercomparison exercise organized during 2007-2008 included six European countries and LBDNet laboratories. Relevant parameters related with dose assessment were evaluated through triage and conventional scoring criteria. A new approach for statistical data analysis was developed including assessment of inter-laboratory reproducibility and intra-laboratory repeatability. Overall, the laboratory performance was satisfactory for mutual cooperation purposes. b) In 2009, LBDNet and two European countries carried out a digital image intercomparison exercise involving dose assessment from metaphase images distributed electronically through internet. The main objectives were to evaluate scoring feasibility on metaphase images and time response. In addition a re-examination phase was considered in which the most controversial images were discussed jointly, this allowed for the development of a homogeneous scoring criteria within the network. c) A further exercise was performed during 2009 involving the shipment of biological samples for biological dosimetry assessment. The aim of this exercise was to test the timely and properly sending and receiving blood samples under national and international regulations. A total of 14 laboratories participated in this joint IAEA, PAHO and WHO. (Author)

  14. NuMA Phosphorylation by Aurora-A Orchestrates Spindle Orientation.

    Science.gov (United States)

    Gallini, Sara; Carminati, Manuel; De Mattia, Fabiola; Pirovano, Laura; Martini, Emanuele; Oldani, Amanda; Asteriti, Italia Anna; Guarguaglini, Giulia; Mapelli, Marina

    2016-02-22

    Spindle positioning is essential for tissue morphogenesis and homeostasis. The signaling network synchronizing spindle placement with mitotic progression relies on timely recruitment at the cell cortex of NuMA:LGN:Gαi complexes, in which NuMA acts as a receptor for the microtubule motor Dynein. To study the implication of Aurora-A in spindle orientation, we developed protocols for the partial inhibition of its activity. Under these conditions, in metaphase NuMA and Dynein accumulate abnormally at the spindle poles and do not reach the cortex, while the cortical distribution of LGN remains unperturbed. FRAP experiments revealed that Aurora-A governs the dynamic exchange between the cytoplasmic and the spindle pole-localized pools of NuMA. We show that Aurora-A phosphorylates directly the C terminus of NuMA on three Ser residues, of which Ser1969 determines the dynamic behavior and the spindle orientation functions of NuMA. Most interestingly, we identify a new microtubule-binding domain of NuMA, which does not overlap with the LGN-binding motif. Our study demonstrates that in metaphase the direct phosphorylation of NuMA by Aurora-A controls its cortical enrichment, and that this is the major event underlying the spindle orientation functions of Aurora-A in transformed and non-transformed cells in culture. Phosphorylation of NuMA by Aurora-A does not affect its affinity for microtubules or for LGN but rather determines the mobility of the protein at the spindle poles. The finding that NuMA can associate concomitantly with LGN and microtubules suggests that its microtubule-binding activity contributes to anchor Dynein-loaded microtubule +TIPs at cortical sites with LGN. Copyright © 2016 Elsevier Ltd. All rights reserved.

  15. Does dietary fat intake influence oocyte competence and embryo quality by inducing oxidative stress in follicular fluid?

    Science.gov (United States)

    Kazemi, Ashraf; Ramezanzadeh, Fatemeh; Nasr-Esfahani, Mohammad Hosein; Saboor Yaraghi, Ali Akbar; Ahmadi, Mehdi

    2013-01-01

    Background: Fat-rich diet may alter oocyte development and maturation and embryonic development by inducing oxidative stress (OS) in follicular environment. Objective: To investigate the relationship between fat intake and oxidative stress with oocyte competence and embryo quality. Materials and Methods: In observational study follicular fluid was collected from 236 women undergoing assisted reproduction program. Malon-di-aldehyde (MDA) levels and total antioxidant capacity (TAC) levels of follicular fluid were assessed as oxidative stress biomarkers. In assisted reproduction treatment cycle fat consumption and its component were assessed. A percentage of metaphase ΙΙ stage oocytes, fertilization rate were considered as markers of oocyte competence and non-fragmented embryo rate, mean of blastomer and good cleavage (embryos with more than 5 cells on 3 days post insemination) rate were considered as markers of embryo quality. Results: The MDA level in follicular fluid was positively related to polyunsaturated fatty acids intake level (p=0.02) and negatively associated with good cleavage rate (p=0.045). Also good cleavage rate (p=0.005) and mean of blastomer (p=0.006) was negatively associated with polyunsaturated fatty acids intake levels. The percentage of metaphase ΙΙ stage oocyte was positively related to the TAC levels in follicular fluid (p=0.046). The relationship between the OS biomarkers in FF and the fertilization rate was not significant. Conclusion: These findings revealed that fat rich diet may induce the OS in oocyte environment and negatively influence embryonic development. This effect can partially be accounted by polyunsaturated fatty acids uptake while oocyte maturation is related to TAC and oocytes with low total antioxidant capacity have lower chance for fertilization and further development. PMID:24639727

  16. Induction of chromosome aberrations and mitotic arrest by cytomegalovirus in human cells

    International Nuclear Information System (INIS)

    AbuBakar, S.; Au, W.W.; Legator, M.S.; Albrecht, T.

    1988-01-01

    Human cytomegalovirus (CMV) is potentially an effective but often overlooked genotoxic agent in humans. We report here evidence that indicates that infection by CMV can induce chromosome alterations and mitotic inhibition. The frequency of chromosome aberrations induced was dependent on the input multiplicity of infection (m.o.i.) for human lung fibroblasts (LU), but not for human peripheral blood lymphocytes (PBLs) when both cell types were infected at the GO phase of the cell cycle. The aberrations induced by CMV were mostly chromatid breaks and chromosome pulverizations that resembled prematurely condensed S-phase chromatin. Pulverized chromosomes were not observed in LU cells infected with virus stocks that had been rendered nonlytic by UV-irradiation at 24,000 ergs/mm2 or from infection of human lymphocytes. In LU cells infected with UV-irradiated CMV, the frequency of aberrations induced was inversely dependent on the extent of the exposure of the CMV stock to the UV-light. In permissive CMV infection of proliferating LU cells at 24 hr after subculture, a high percentage (greater than 40%) of the metaphase cells were arrested at their first metaphase and displayed severely condensed chromosomes when harvested 48 hr later. A significant increase (p less than 0.05) in the chromosome aberration frequency was also observed. Our study shows that CMV infection is genotoxic to host cells. The types and extent of damage are dependent on the viral genome expression and on the cell cycle stage of the cells at the time of infection. The possible mechanisms for induction of chromosome damage by CMV are discussed

  17. The Drosophila orthologue of the INT6 onco-protein regulates mitotic microtubule growth and kinetochore structure.

    Directory of Open Access Journals (Sweden)

    Fioranna Renda

    2017-05-01

    Full Text Available INT6/eIF3e is a highly conserved component of the translation initiation complex that interacts with both the 26S proteasome and the COP9 signalosome, two complexes implicated in ubiquitin-mediated protein degradation. The INT6 gene was originally identified as the insertion site of the mouse mammary tumor virus (MMTV, and later shown to be involved in human tumorigenesis. Here we show that depletion of the Drosophila orthologue of INT6 (Int6 results in short mitotic spindles and deformed centromeres and kinetochores with low intra-kinetochore distance. Poleward flux of microtubule subunits during metaphase is reduced, although fluorescence recovery after photobleaching (FRAP demonstrates that microtubules remain dynamic both near the kinetochores and at spindle poles. Mitotic progression is delayed during metaphase due to the activity of the spindle assembly checkpoint (SAC. Interestingly, a deubiquitinated form of the kinesin Klp67A (a putative orthologue of human Kif18A accumulates near the kinetochores in Int6-depleted cells. Consistent with this finding, Klp67A overexpression mimics the Int6 RNAi phenotype. Furthermore, simultaneous depletion of Int6 and Klp67A results in a phenotype identical to RNAi of just Klp67A, which indicates that Klp67A deficiency is epistatic over Int6 deficiency. We propose that Int6-mediated ubiquitination is required to control the activity of Klp67A. In the absence of this control, excess of Klp67A at the kinetochore suppresses microtubule plus-end polymerization, which in turn results in reduced microtubule flux, spindle shortening, and centromere/kinetochore deformation.

  18. Chromosomal assignment of canine THADA gene to CFA 10q25

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    Dolf Gaudenz

    2008-06-01

    Full Text Available Abstract Background Chromosomal translocations affecting the chromosome 2p21 cluster in a 450 kb breakpoint region are frequently observed in human benign thyroid adenomas. THADA (thyroid adenoma associated was identified as the affected gene within this breakpoint region. In contrast to man tumours of the thyroid gland of dogs (Canis lupus familiaris constitute mainly as follicular cell carcinomas, with malignant thyroid tumours being more frequent than benign thyroid adenomas. In order to elucidate if the THADA gene is also a target of chromosomal rearrangements in thyroid adenomas of the dog we have physically mapped the canine THADA gene to canine chromosome 10. A PCR was established to screen a canine genome library for a BAC clone containing the gene sequence of canine THADA. Further PCR reactions were done using the identified BAC clone as a template in order to verify the corresponding PCR product by sequencing. Canine whole blood was incubated with colcemid in order to arrest the cultured cells in metaphases. The verified BAC DNA was digoxigenin labeled and used as a probe in fluorescence in situ hybridization (FISH. Ten well spread metaphases were examined indicating a signal on canine chromosome 10 on both chromatids. A detailed fine mapping was performed indicating the canine THADA gene locus on the q-arm of chromosome 10. Results The canine THADA gene locus was mapped on chromosome 10q25. Our mapping results obtained in this study following the previously described nomenclature for the canine karyotype. Conclusion We analysed whether the THADA gene locus is a hotspot of canine chromosomal rearrangements in canine neoplastic lesions of the thyroid and in addition might play a role as a candidate gene for a possible malignant transformation of canine thyroid adenomas. Although the available cytogenetic data of canine thyroid adenomas are still insufficient the chromosomal region to which the canine THADA has been mapped seems to be no

  19. Cdc20 is critical for meiosis I and fertility of female mice.

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    Fang Jin

    2010-09-01

    Full Text Available Chromosome missegregation in germ cells is an important cause of unexplained infertility, miscarriages, and congenital birth defects in humans. However, the molecular defects that lead to production of aneuploid gametes are largely unknown. Cdc20, the activating subunit of the anaphase-promoting complex/cyclosome (APC/C, initiates sister-chromatid separation by ordering the destruction of two key anaphase inhibitors, cyclin B1 and securin, at the transition from metaphase to anaphase. The physiological significance and full repertoire of functions of mammalian Cdc20 are unclear at present, mainly because of the essential nature of this protein in cell cycle progression. To bypass this problem we generated hypomorphic mice that express low amounts of Cdc20. These mice are healthy and have a normal lifespan, but females produce either no or very few offspring, despite normal folliculogenesis and fertilization rates. When mated with wild-type males, hypomorphic females yield nearly normal numbers of fertilized eggs, but as these embryos develop, they become malformed and rarely reach the blastocyst stage. In exploring the underlying mechanism, we uncover that the vast majority of these embryos have abnormal chromosome numbers, primarily due to chromosome lagging and chromosome misalignment during meiosis I in the oocyte. Furthermore, cyclin B1, cyclin A2, and securin are inefficiently degraded in metaphase I; and anaphase I onset is markedly delayed. These results demonstrate that the physiologically effective threshold level of Cdc20 is high for female meiosis I and identify Cdc20 hypomorphism as a mechanism for chromosome missegregation and formation of aneuploid gametes.

  20. Dose-response curve for blood exposed to gamma-neutron mixed field by conventional cytogenetic method

    International Nuclear Information System (INIS)

    Brandao, Jose Odinilson de C.; Souza, Priscilla L.G.; Santos, Joelan A.L.; Vilela, Eudice C.; Lima, Fabiana F.; Calixto, Merilane S.; Santos, Neide

    2009-01-01

    There is increasing concern about airline crew members (about one million worldwide) are exposed to measurable neutrons doses. Historically, cytogenetic biodosimetry assays have been based on quantifying asymmetrical chromosome alterations (dicentrics, centric rings and acentric fragments) in mytogen-stimulated T-lymphocytes in their first mitosis after radiation exposure. Increased levels of chromosome damage in peripheral blood lymphocytes are a sensitive indicator of radiation exposure and they are routinely exploited for assessing radiation absorbed dose after accidental or occupational exposure. Since radiological accidents are not common, not all nations feel that it is economically justified to maintain biodosimetry competence. However, dependable access to biological dosimetry capabilities is completely critical in event of an accident. In this paper the dose-response curve was measured for the induction of chromosomal alterations in peripheral blood lymphocytes after chronic exposure in vitro to neutron-gamma mixes field. Blood was obtained from one healthy donor and exposed to two neutron-gamma mixed field from sources 241 AmBe (20 Ci) at the Neutron Calibration Laboratory (NCL-CRCN/NE-PE-Brazil). The evaluated absorbed doses were 0.2 Gy; 1.0 Gy and 2.5 Gy. The dicentric chromosomes were observed at metaphase, following colcemid accumulation and 1000 well-spread metaphase figures were analyzed for the presence of dicentrics by two experienced scorers after painted by giemsa 5%. Our preliminary results showed a linear dependence between radiations absorbed dose and dicentric chromosomes frequencies. Dose-response curve described in this paper will contribute to the construction of calibration curve that will be used in our laboratory for biological dosimetry. (author)

  1. Chromosomal Fragmentation: A Possible Marker for the Selection of High Gymnemic Acid Yielding Accessions ofGymnema sylvestreR. Br.

    Science.gov (United States)

    Verma, Ashutosh Kumar; Dhawan, Sunita Singh

    2017-10-01

    Gymnema sylvestre R. Br. a member of family Asclepiadaceae as mentioned in Indian Pharmacopoeia popular among the researchers because of stimulatory effect of its phytoconstituent on pancreatic cells and potential to treat Type I and II type of diabetes. Development of cost-effective marker system for the selection of high gymnemic acid yielding accessions of G. sylvestre . Presoaked seeds of Brassica campestris treated with different dilutions of gymnemagenin and 10% leaf extract of twenty different accessions of G. sylvestre . Root tips of germinated seeds were fixed, and chromosomal studies were made by root tip bioassay method. Exposure of seeds to treatment solutions promotes various types of chromosomal anomalies in root meristem, and surprisingly, direct correlation between the percentage of chromosomal fragmentation and the percentage of gymnemic acid shared by treatment solution were observed. Later finding may be explored for the development of a novel methodology or marker system for the selection of high active principle yielding accessions of G. sylvestre . An experiment was carried out using root tip bioassay method for the study of effect of different dilutions of standard gymnemic acid and 10% leaf extract of twenty different accessions of Gymnema sylvestre on root tip meristem of Brassica campestris . Various types of chromosomal anomalies were observed. Of which, percentage of chromosomal fragmentation was showed a direct (∞) relationship with the percentage of gymnemic acid shared by treatment solution. This interesting result after more and more exploration and revalidation could be utilized for the development of a novel methodology for the selection of high active principle yielding accessions of G. sylvestre . Abbreviations used: MI: Mitotic index; CP: Condensed prophase; CM: Clumped metaphase; MC: Metaphase cleft; FR: Fragmentation; AP: Anaphase with persistent nucleolous; LA: Laggard, BR: Bridge; BI: Bi-nucleated cell; DA: Disturbed

  2. The GnRH analogue triptorelin confers ovarian radio-protection to adult female rats

    Energy Technology Data Exchange (ETDEWEB)

    Camats, N. [Institut de Biotecnologia i de Biomedicina (I.B.B.), Universitat Autonoma de Barcelona, 08193 Barcelona (Spain); Departament de Biologia Cel.lular, Fisiologia i Immunologia, Universitat Autonoma de Barcelona, 08193 Barcelona (Spain); Garcia, F. [Institut de Biotecnologia i de Biomedicina (I.B.B.), Universitat Autonoma de Barcelona, 08193 Barcelona (Spain); Parrilla, J.J. [Servicio de Ginecologia y Obstetricia, Hospital Universitario Virgen de la Arrixaca, 30120 El Palmar, Murcia (Spain); Calaf, J. [Servei de Ginecologia i Obstetricia, Hospital Universitari de la Santa Creu i Sant Pau, 08025 Barcelona (Spain); Martin-Mateo, M. [Departament de Pediatria, d' Obstetricia i Ginecologia i de Medicina Preventiva, Universitat Autonoma de Barcelona, 08193 Barcelona (Spain); Caldes, M. Garcia, E-mail: Montserrat.Garcia.Caldes@uab.es [Institut de Biotecnologia i de Biomedicina (I.B.B.), Universitat Autonoma de Barcelona, 08193 Barcelona (Spain); Departament de Biologia Cel.lular, Fisiologia i Immunologia, Universitat Autonoma de Barcelona, 08193 Barcelona (Spain)

    2009-10-02

    There is a controversy regarding the effects of the analogues of the gonadotrophin-releasing hormone (GnRH) in radiotherapy. This has led us to study the possible radio-protection of the ovarian function of a GnRH agonist analogue (GnRHa), triptorelin, in adult, female rats (Rattus norvegicus sp.). The effects of the X-irradiation on the oocytes of ovarian primordial follicles, with and without GnRHa treatment, were compared, directly in the female rats (F{sub 0}) with reproductive parameters, and in the somatic cells of the resulting foetuses (F{sub 1}) with cytogenetical parameters. In order to do this, the ovaries and uteri from 82 females were extracted for the reproductive analysis and 236 foetuses were obtained for cytogenetical analysis. The cytogenetical study was based on the data from 22,151 metaphases analysed. The cytogenetical parameters analysed to assess the existence of chromosomal instability were the number of aberrant metaphases (2234) and the number (2854) and type of structural chromosomal aberrations, including gaps and breaks. Concerning the reproductive analysis of the ovaries and the uteri, the parameters analysed were the number of corpora lutea, implantations, implantation losses and foetuses. Triptorelin confers radio-protection of the ovaries in front of chromosomal instability, which is different, with respect to the single and fractioned dose. The cytogenetical analysis shows a general decrease in most of the parameters of the triptorelin-treated groups, with respect to their controls, and some of these differences were considered to be statistically significant. The reproductive analysis indicates that there is also radio-protection by the agonist, although minor to the cytogenetical one. Only some of the analysed parameters show a statistically significant decrease in the triptorelin-treated groups.

  3. Analysis of chromosomal abnormalities: a study of partial exposure to X-rays

    International Nuclear Information System (INIS)

    Andrade, Aida M.G. de; Mendes, Mariana E.; Mendonça, Julyanne C.G.; Melo, Laís; Hwang, Suy; Santos, Neide; Lima, Fabiana F. de; Centro Regional de Ciências Nucleares; Universidade Federal de Pernambuco

    2017-01-01

    Biological dosimetry is used in case of supposed accidental overexposure. The most commonly used biomarkers for assessing the absorbed dose are unstable chromosomal abnormalities. In a case of a partial body exposure, the frequencies of those abnormalities varies according to the area of the exposed body and may be substantially different from a total exposure of the body with an identical dose. The present study aimed to evaluate the frequency of chromosomal changes simulating, with blood samples, partial (25%, 50%) and full body irradiation (100%) in X-ray beam. The irradiation was performed at Metrology Service (CRCN-NE / CNEN) with a bundle of 250kVp X-rays, resulting in the absorbed dose of 1.0 Gy. Prior to obtain the metaphases, irradiated blood was mixed with non-irradiated blood, and then the mitotic metaphases for the chromosomal analyzes were obtained by culturing lymphocytes and the slides were stained with 5% Giemsa. It was observed that there was an increase in dicentric frequency when the dose percentage increases in both subjects (0.024 and 0.049 in subject 1 and 0.016 and 0.038 in subject 2) after irradiation. The cellular distribution was 'contaminated' only at dose 25% of the first individual who had a prolongation of the distribution. The Qdr and Dolphin methods were used to estimate partial absorbed dose, but the Qdr method was not efficient and whereas the Dolphin method was efficient when the individual had a prolonged cell distribution. It is necessary to increase the number of observations to be sure of the observed behaviors. (author)

  4. Kinetochore-microtubule attachment throughout mitosis potentiated by the elongated stalk of the kinetochore kinesin CENP-E.

    Science.gov (United States)

    Vitre, Benjamin; Gudimchuk, Nikita; Borda, Ranier; Kim, Yumi; Heuser, John E; Cleveland, Don W; Grishchuk, Ekaterina L

    2014-08-01

    Centromere protein E (CENP-E) is a highly elongated kinesin that transports pole-proximal chromosomes during congression in prometaphase. During metaphase, it facilitates kinetochore-microtubule end-on attachment required to achieve and maintain chromosome alignment. In vitro CENP-E can walk processively along microtubule tracks and follow both growing and shrinking microtubule plus ends. Neither the CENP-E-dependent transport along microtubules nor its tip-tracking activity requires the unusually long coiled-coil stalk of CENP-E. The biological role for the CENP-E stalk has now been identified through creation of "Bonsai" CENP-E with significantly shortened stalk but wild-type motor and tail domains. We demonstrate that Bonsai CENP-E fails to bind microtubules in vitro unless a cargo is contemporaneously bound via its C-terminal tail. In contrast, both full-length and truncated CENP-E that has no stalk and tail exhibit robust motility with and without cargo binding, highlighting the importance of CENP-E stalk for its activity. Correspondingly, kinetochore attachment to microtubule ends is shown to be disrupted in cells whose CENP-E has a shortened stalk, thereby producing chromosome misalignment in metaphase and lagging chromosomes during anaphase. Together these findings establish an unexpected role of CENP-E elongated stalk in ensuring stability of kinetochore-microtubule attachments during chromosome congression and segregation. © 2014 Vitre, Gudimchuk, et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

  5. The extracellular matrix of porcine mature oocytes: Origin, composition and presumptive roles

    Directory of Open Access Journals (Sweden)

    Pivko Juraj

    2003-12-01

    Full Text Available Abstract The extracellular matrix (ECM of porcine mature oocytes was revealed by transmission electron microscopy (TEM after treatment with tannic acid and ruthenium red. Present in the perivitelline space (PVS and on the surface of the zona pellucida (ZP, it appeared to be composed of thin filaments and granules at the interconnections of the filaments, which were interpreted respectively as hyaluronic acid chains and bound proteoglycans. In order to determine whether this material is produced by the corona cells (the same ECM was found also on the surface of the zona pellucida and between cumulus cells or by the oocyte itself, the synthesis of glycoproteins and glycosaminoglycans was checked by autoradiography on semi-thin and thin sections observed by light and electron microscopy. Immature oocytes within or without cumulus cells, were incubated with L [3H-] fucose or L [3H-] glucosamine – precursors respectively of glycoproteins and hyaluronic acid or hyaluronan (HA bound to proteoglycans – for various times (with or without chase and at different stages during in vitro maturation. In the first case, incorporation was found in both cumulus cells and ooplasm (notably in the Golgi area for 3H-fucose and labeled material accumulated in the ECM of the PVS and of the ZP surface. Labeling in the PVS with both precursors was maximum between metaphase I (MI and metaphase II (MII and was partially extracted by hyaluronidase but not by neuraminidase. Tunicamycin, an inhibitor of glycoprotein synthesis, significantly decreased the amount of 3H-fucose labeled molecules in the PVS and increased the incidence of polyspermic penetration during subsequent in vivo fertilization. Since cumulus-free oocytes also secreted 3H-glucosamine containing compounds, both oocyte and cumulus cells probably contribute to the production of the ECM found in the PVS of mature oocytes. ECM and particularly its HA moiety present on both sides of the ZP may constitute a

  6. Expression pattern of G protein‑coupled estrogen receptor 1 (GPER) in human cumulus granulosa cells (CGCs) of patients with PCOS.

    Science.gov (United States)

    Zang, Lili; Zhang, Quan; Zhou, Yi; Zhao, Yan; Lu, Linlin; Jiang, Zhou; Peng, Zhen; Zou, Shuhua

    2016-06-01

    Estradiol mediates its actions by binding to classical nuclear receptors, estrogen receptor α (ER-α) and estrogen receptor β (ER-β), and the non-classical G protein-coupled estrogen receptor 1(GPER). Several gene knockdown models have shown the importance of the receptors for growth of the oocyte and for ovulation. The aim of our study was to identify the pattern of GPER expression in human cumulus granulosa cells (CGCs) from ovarian follicles at different stages of oocyte maturation, and the differences of GPER expression between polycystic ovary syndrome (PCOS) patients and non-PCOS women. Thirty-eight cases of PCOS patients and a control group of thirty-two infertile women without PCOS were used in this study. GPER's location in CGCs was investigated by immunohistochemistry. Quantitative RT-PCR and western blot were used to identify the quantify GPER expression. Here we demonstrated that GPER was expressed in CGCs of both PCOS patients and non-PCOS women, and the expression of GPER was decreased significantly during oocyte maturation. But the expression levels of GPER in CGCs of PCOS patients and non-PCOS women were not significantly different. The data indicate that GPER may play a role during human oocyte maturation through its action in cumulus granulosa cells. AMHRIIs: anti-Mullerian hormone type II receptors; BMI: body mass index; CGCs: cumulus granulosa cells; COH: controlled ovarian hyperstimulation; E2: estradiol; EGFR: epidermal growth factor receptor; ER-α: estrogen receptor; ER-β: estrogen receptor β; FF: follicular fluid; FSH: follicle-stimulating hormone; GCs: granulosa cells; GPER: G protein-coupled estrogen receptor 1; GV: germinal vesicle; GVBD: germinal vesicle breakdown; HCG: human chorionic gonadotropin; IRS: immunoreactive score; IVF-ET: in vitro fertilization and embryo transfer; MI: metaphase I; MII: metaphase II; MAPK: mitogen-activated protein kinase; OCCCs: oocyte corona cumulus complexes; PCOS: polycystic ovarian syndrome; q

  7. Heterogeneous nuclear ribonucleoprotein K upregulates the kinetochore complex component NUF2 and promotes the tumorigenicity of colon cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Sugimasa, Hironobu; Taniue, Kenzui [Laboratory of Molecular and Genetic Information, Institute of Molecular and Cellular Biosciences, The University of Tokyo, 1-1-1, Yayoi, Bunkyo-ku, Tokyo, 113-0032 (Japan); Kurimoto, Akiko [Laboratory of Molecular and Genetic Information, Institute of Molecular and Cellular Biosciences, The University of Tokyo, 1-1-1, Yayoi, Bunkyo-ku, Tokyo, 113-0032 (Japan); Oncology Research Laboratories, Daiichi Sankyo Co., Ltd, 1-2-58, Hiromachi, Shinagawa-ku, Tokyo, 140-8710 (Japan); Takeda, Yasuko; Kawasaki, Yoshihiro [Laboratory of Molecular and Genetic Information, Institute of Molecular and Cellular Biosciences, The University of Tokyo, 1-1-1, Yayoi, Bunkyo-ku, Tokyo, 113-0032 (Japan); Akiyama, Tetsu, E-mail: akiyama@iam.u-tokyo.ac.jp [Laboratory of Molecular and Genetic Information, Institute of Molecular and Cellular Biosciences, The University of Tokyo, 1-1-1, Yayoi, Bunkyo-ku, Tokyo, 113-0032 (Japan)

    2015-03-27

    Heterogeneous nuclear ribonucleoprotein K (hnRNP K) is a multi-functional protein involved in transcription, mRNA splicing, mRNA stabilization and translation. Although hnRNP K has been suggested to play a role in the development of many cancers, its molecular function in colorectal cancer has remained elusive. Here we show that hnRNP K plays an important role in the mitotic process in HCT116 colon cancer cells. Furthermore, we demonstrate that hnRNP K directly transactivates the NUF2 gene, the product of which is a component of the NDC80 kinetochore complex and which is known to be critical for a stable spindle microtubule-kinetochore attachment. In addition, knockdown of both hnRNP K and NUF2 caused failure in metaphase chromosome alignment and drastic decrease in the growth of colon cancer cells. These results suggest that the hnRNP K-NUF2 axis is important for the mitotic process and proliferation of colon cancer cells and that this axis could be a target for the therapy of colon cancer. - Highlights: • hnRNP K is required for the tumorigenicity of colon cancer cells. • hnRNP K binds to the promoter region of NUF2 and activates its transcription. • NUF2 expression is correlated with hnRNP K expression in colorectal cancer tissue. • hnRNP K and NUF2 are required for metaphase chromosome alignment. • The hnRNP K-NUF2 axis is important for the proliferation of colon cancer cells.

  8. v-Src causes delocalization of Mklp1, Aurora B, and INCENP from the spindle midzone during cytokinesis failure

    Energy Technology Data Exchange (ETDEWEB)

    Soeda, Shuhei [Department of Molecular Cell Biology, Graduate School of Pharmaceutical Sciences, Chiba University, Inohana 1-8-1, Chuo-ku, Chiba 260-8675 (Japan); Nakayama, Yuji, E-mail: nakayama@mb.kyoto-phu.ac.jp [Department of Molecular Cell Biology, Graduate School of Pharmaceutical Sciences, Chiba University, Inohana 1-8-1, Chuo-ku, Chiba 260-8675 (Japan); Department of Biochemistry and Molecular Biology, Kyoto Pharmaceutical University, 5 Nakauchi-cho, Misasagi, Yamashina-ku, Kyoto 607-8414 (Japan); Honda, Takuya; Aoki, Azumi; Tamura, Naoki; Abe, Kohei; Fukumoto, Yasunori [Department of Molecular Cell Biology, Graduate School of Pharmaceutical Sciences, Chiba University, Inohana 1-8-1, Chuo-ku, Chiba 260-8675 (Japan); Yamaguchi, Naoto, E-mail: nyama@faculty.chiba-u.jp [Department of Molecular Cell Biology, Graduate School of Pharmaceutical Sciences, Chiba University, Inohana 1-8-1, Chuo-ku, Chiba 260-8675 (Japan)

    2013-06-10

    Src-family tyrosine kinases are aberrantly activated in cancers, and this activation is associated with malignant tumor progression. v-Src, encoded by the v-src transforming gene of the Rous sarcoma virus, is a mutant variant of the cellular proto-oncogene c-Src. Although investigations with temperature sensitive mutants of v-Src have shown that v-Src induces many oncogenic processes, the effects on cell division are unknown. Here, we show that v-Src inhibits cellular proliferation of HCT116, HeLa S3 and NIH3T3 cells. Flow cytometry analysis indicated that inducible expression of v-Src results in an accumulation of 4N cells. Time-lapse analysis revealed that binucleation is induced through the inhibition of cytokinesis, a final step of cell division. The localization of Mklp1, which is essential for cytokinesis, to the spindle midzone is inhibited in v-Src-expressing cells. Intriguingly, Aurora B, which regulates Mklp1 localization at the midzone, is delocalized from the spindle midzone and the midbody but not from the metaphase chromosomes upon v-Src expression. Mklp2, which is responsible for the relocation of Aurora B from the metaphase chromosomes to the spindle midzone, is also lost from the spindle midzone. These results suggest that v-Src inhibits cytokinesis through the delocalization of Mklp1 and Aurora B from the spindle midzone, resulting in binucleation. -- Highlights: • v-Src inhibits cell proliferation of HCT116, HeLa S3 and NIH3T3 cells. • v-Src induces binucleation together with cytokinesis failure. • v-Src causes delocalization of Mklp1, Aurora B and INCENP from the spindle midzone.

  9. Clinical and biological significance of isolated Y chromosome loss in myelodysplastic syndromes and chronic myelomonocytic leukemia. A report from the Spanish MDS Group.

    Science.gov (United States)

    Nomdedeu, Meritxell; Pereira, Arturo; Calvo, Xavier; Colomer, Joan; Sole, Francesc; Arias, Amparo; Gomez, Candida; Luño, Elisa; Cervera, Jose; Arnan, Montserrat; Pomares, Helena; Ramos, Fernando; Oiartzabal, Itziar; Espinet, Blanca; Pedro, Carme; Arrizabalaga, Beatriz; Blanco, María Laura; Tormo, Mar; Hernandez-Rivas, Jesus Maria; Díez-Campelo, María; Ortega, Margarita; Valcárcel, David; Cedena, Maria-Teresa; Collado, Rosa; Grau, Javier; Granada, Isabel; Sanz, Guillermo; Campo, Elias; Esteve, Jordi; Costa, Dolors

    2017-12-01

    Isolate loss of chromosome Y (-Y) in myelodysplastic syndromes (MDS) is associated to a better outcome but it is also well described as an age-related phenomenon. In this study we aimed to analyze the prognostic impact of -Y in the context of the IPSS-R cytogenetic classification, evaluate the clinical significance of the percentage of metaphases with isolated -Y, and test whether finding -Y may predispose to over-diagnose MDS in patients with borderline morphological features. We evaluated 3581 male patients from the Spanish MDS Registry with a diagnosis of MDS or chronic myelomonocytic leukemia (CMML). -Y was identified in 177 patients (4.9%). Compared with the 2246 male patients with normal karyotype, -Y group showed a reduced risk of leukemic transformation that did not translate into a survival advantage. The overall survival and the risk of leukemic transformation were not influenced by the percentage of metaphases with -Y. The -Y group was not enriched in patients with minor morphologic traits of dysplasia, suggesting that the better outcome in the -Y group cannot be explained by enrichment in cases misdiagnosed as MDS. In conclusion, our results support the current recommendation of classifying patients with -Y within the very good risk category of the IPSS-R for MDS and rule out a selection bias as a possible explanation of this better outcome. An analysis of the molecular basis of MDS with isolated -Y would be of interest as it may provide a biological basis of protection against progression to acute leukemia. Copyright © 2017 Elsevier Ltd. All rights reserved.

  10. Optimized Fast-FISH with a-satellite probes: acceleration by microwave activation

    Directory of Open Access Journals (Sweden)

    Durm M.

    1997-01-01

    Full Text Available It has been shown for several DNA probes that the recently introduced Fast-FISH (fluorescence in situ hybridization technique is well suited for quantitative microscopy. For highly repetitive DNA probes the hybridization (renaturation time and the number of subsequent washing steps were reduced considerably by omitting denaturing chemical agents (e.g., formamide. The appropriate hybridization temperature and time allow a clear discrimination between major and minor binding sites by quantitative fluorescence microscopy. The well-defined physical conditions for hybridization permit automatization of the procedure, e.g., by a programmable thermal cycler. Here, we present optimized conditions for a commercially available X-specific a-satellite probe. Highly fluorescent major binding sites were obtained for 74oC hybridization temperature and 60 min hybridization time. They were clearly discriminated from some low fluorescent minor binding sites on metaphase chromosomes as well as in interphase cell nuclei. On average, a total of 3.43 ± 1.59 binding sites were measured in metaphase spreads, and 2.69 ± 1.00 in interphase nuclei. Microwave activation for denaturation and hybridization was tested to accelerate the procedure. The slides with the target material and the hybridization buffer were placed in a standard microwave oven. After denaturation for 20 s at 900 W, hybridization was performed for 4 min at 90 W. The suitability of a microwave oven for Fast-FISH was confirmed by the application to a chromosome 1-specific a-satellite probe. In this case, denaturation was performed at 630 W for 60 s and hybridization at 90 W for 5 min. In all cases, the results were analyzed quantitatively and compared to the results obtained by Fast-FISH. The major binding sites were clearly discriminated by their brightness

  11. Single, repeated dose toxicity and genotoxicity assessment of herb formula KIOM2012H

    Directory of Open Access Journals (Sweden)

    Hwayong Park

    2017-12-01

    Full Text Available Background: Traditional medicine and herbal prescriptions are becoming more popular, and they account for a large share of the world’s healthcare research studies, developments, and market demands. Increasing scientific evidence of the substantive efficacies such as preventive health keeping pharmaceutical materials and dietary supplements can be found elsewhere. Above all, safety should be the critical premise for considering developmental materials such as pharmaceuticals without side effects and toxicity. Methods: The authors formulated KIOM2012H (K2H using four herbs that were reported to have medicinal effects—including anticancer, antiaging, antimicrobial, inflammation, and neuroprotective properties. In order to examine the toxicity, single and repeated dose toxicity, and genotoxicities of bacterial mutation, micronucleus, and chromosomal aberration assays were conducted. Results: All experimental observations and results showed normal findings. Toxicities or abnormal signs were not observed in all experimental assays, including oral administration, animal behavior, clinical findings, and changes in body weight in vivo. In vitro bacterial cultures produced no revertant colonies, and no increased numbers of structural or numerical aberrant metaphases were found in the metaphase chromosomes examined. Moreover, no significant increased frequency of micronucleus was observed in any of the doses used. Overall, no acute toxicity or genotoxicity was found in all analysis parameters in all the assays conducted. Conclusion: Reviewing the results as a whole, K2H extract was regarded as a safe material with no toxicity, and can be applied for the research and development of complementary and alternative medicines with improved efficacy in current therapeutic healthcare, based on traditional medicine and herb resources. Keywords: genotoxicity, herb, toxicity, traditional medicine

  12. Heterogeneous nuclear ribonucleoprotein K upregulates the kinetochore complex component NUF2 and promotes the tumorigenicity of colon cancer cells

    International Nuclear Information System (INIS)

    Sugimasa, Hironobu; Taniue, Kenzui; Kurimoto, Akiko; Takeda, Yasuko; Kawasaki, Yoshihiro; Akiyama, Tetsu

    2015-01-01

    Heterogeneous nuclear ribonucleoprotein K (hnRNP K) is a multi-functional protein involved in transcription, mRNA splicing, mRNA stabilization and translation. Although hnRNP K has been suggested to play a role in the development of many cancers, its molecular function in colorectal cancer has remained elusive. Here we show that hnRNP K plays an important role in the mitotic process in HCT116 colon cancer cells. Furthermore, we demonstrate that hnRNP K directly transactivates the NUF2 gene, the product of which is a component of the NDC80 kinetochore complex and which is known to be critical for a stable spindle microtubule-kinetochore attachment. In addition, knockdown of both hnRNP K and NUF2 caused failure in metaphase chromosome alignment and drastic decrease in the growth of colon cancer cells. These results suggest that the hnRNP K-NUF2 axis is important for the mitotic process and proliferation of colon cancer cells and that this axis could be a target for the therapy of colon cancer. - Highlights: • hnRNP K is required for the tumorigenicity of colon cancer cells. • hnRNP K binds to the promoter region of NUF2 and activates its transcription. • NUF2 expression is correlated with hnRNP K expression in colorectal cancer tissue. • hnRNP K and NUF2 are required for metaphase chromosome alignment. • The hnRNP K-NUF2 axis is important for the proliferation of colon cancer cells

  13. Evaluation of the clastogenicity and anticlastogenicity of the carotenoid bixin in human lymphocyte cultures.

    Science.gov (United States)

    Antunes, Lusânia M Greggi; Pascoal, Lívia M; Bianchi, Maria de Lourdes P; Dias, Francisca L

    2005-08-01

    Carotenoids are regarded as effective antioxidants, antimutagenic and anticarcinogenic agents. Annatto, a red-yellow extract obtained from seeds of Bixa orellana L. is a mixture of several carotenoids and one of them bixin (BXN), is known as its major coloring compound. Studies on BXN clastogenicity and anticlastogenicity in cultured human lymphocytes have not been reported so far. Therefore, the present study was undertaken to investigate the ability of BXN to induce chromosomal aberrations in human lymphocytes in vitro and to examine the possible anticlastogenic effect of this carotenoid in chromosomal damage induced by the clastogen cisplatin (cDDP). Human blood samples were obtained from six healthy, non-smoking volunteers; two females and four males aged 18-35 years. The concentrations of BXN (1.0; 2.5; 5.0 or 10 microg/mL) tested in combination with cDDP were established on the basis of mitotic index (MI) measurements. The data showed that BXN was not cytotoxic or clastogenic, when compared to untreated control. A marked decrease in the MI values compared to the untreated control and an increased percentage of aberrant metaphases was seen in all cultures treated with cDDP. The carotenoid efficiency in reducing the inhibitory effect of cDDP on lymphocyte MI is concentration-dependent. Cultures simultaneously treated with BXN and cDDP showed a statistically significant reduction in total chromosomal aberrations and aberrant metaphases. In our experiments, BXN may have acted as an antioxidant by intercepting free radicals generated by cDDP. The data obtained in the present study suggest that dietary carotenoids may act as protective agents against clastogenic effects of antitumor agents. However, extensive studies are necessary to elucidate the mechanism of action of BXN before its therapeutic use.

  14. Detection of 16 p deletions by FISH in patients with inv(16) or t(16;16) and acute myeloid leukemia (AML).

    Science.gov (United States)

    Martinet, D; Mühlematter, D; Leeman, M; Parlier, V; Hess, U; Gmür, J; Jotterand, M

    1997-07-01

    Deletions of sequences centromeric to the p-arm breakpoint have been described in a subset of patients with inv(16) and acute myeloid leukemia (AML) and reported to be associated with a relatively good prognosis. We have investigated 16 p deletions in a cohort of 15 patients with AML and inv(16) or t(16;16) and compared non-deletion and deletion patients in terms of clinical course. Patients were studied by fluorescence in situ hybridization (FISH) using cosmid zit14 as a probe to detect the presence of 16 p deletions in metaphase chromosomes of leukemic cells. While seven patients (47%) revealed no evidence of a deletion, five patients (33%) presented 16 p deletions, thus bringing further support to the relatively frequent occurrence of this event in inv(16) patients. Remarkably, two patients with inv(16) and one patient with t(16;16) showed a mosaicism of deletion and non-deletion metaphases suggesting the presence of two distinct leukemic cell populations. Results let us assume that 16 p deletions are not restricted to inv(16) and may occur subsequently to inv(16) or t(16;16). The presence of a 16 p deletion in a case of inv(16) associated with CBFB-MYH11 transcript type E indicates that deletions are not limited to CBFB-MYH11 transcript type A rearrangements. Survival of deletion patients was compared with that of non-deletion and mosaic ones. No significant differences were observed. The advantage of FISH for enumerative and quantitative assessment of submicroscopic rearrangements of clinical significance is further emphasized.

  15. Karyotype of cryopreserved bone marrow cells

    Directory of Open Access Journals (Sweden)

    M.L.L.F. Chauffaille

    2003-07-01

    Full Text Available The analysis of chromosomal abnormalities is important for the study of hematological neoplastic disorders since it facilitates classification of the disease. The ability to perform chromosome analysis of cryopreserved malignant marrow or peripheral blast cells is important for retrospective studies. In the present study, we compared the karyotype of fresh bone marrow cells (20 metaphases to that of cells stored with a simplified cryopreservation method, evaluated the effect of the use of granulocyte-macrophage colony-stimulating factor (GM-CSF as an in vitro mitotic index stimulator, and compared the cell viability and chromosome morphology of fresh and cryopreserved cells whenever possible (sufficient metaphases for analysis. Twenty-five bone marrow samples from 24 patients with hematological disorders such as acute myeloid leukemia, acute lymphoblastic leukemia, myelodysplastic syndrome, chronic myeloid leukemia, megaloblastic anemia and lymphoma (8, 3, 3, 8, 1, and 1 patients, respectively were selected at diagnosis, at relapse or during routine follow-up and one sample was obtained from a bone marrow donor after informed consent. Average cell viability before and after freezing was 98.8 and 78.5%, respectively (P < 0.05. Cytogenetic analysis was successful in 76% of fresh cell cultures, as opposed to 52% of cryopreserved samples (P < 0.05. GM-CSF had no proliferative effect before or after freezing. The morphological aspects of the chromosomes in fresh and cryopreserved cells were subjectively the same. The present study shows that cytogenetic analysis of cryopreserved bone marrow cells can be a reliable alternative when fresh cell analysis cannot be done, notwithstanding the reduced viability and lower percent of successful analysis that are associated with freezing.

  16. Alteration of energy metabolism gene expression in cumulus cells affects oocyte maturation via MOS-mitogen-activated protein kinase pathway in dairy cows with an unfavorable "Fertil-" haplotype of one female fertility quantitative trait locus.

    Science.gov (United States)

    Brisard, Daphné; Desmarchais, Alice; Touzé, Jean-Luc; Lardic, Lionel; Freret, Sandrine; Elis, Sebastien; Nuttinck, Fabienne; Camous, Sylvaine; Dupont, Joelle; Uzbekova, Svetlana

    2014-03-01

    Prim'Holstein heifers selected for the "Fertil-" homozygous haplotype of QTL-Female-Fert ility-BTA3 showed a greater rate of early pregnancy failure and slower embryo development after IVM suggesting lower oocyte quality than those selected for "Fertile+". We aimed to ascertain intrafollicular factors related to lower oocyte quality in "Fertil-" cows. Analysis of individual oocytes showed meiotic progression delay in "Fertil-" compared with "Fertil+" dairy cows after in vivo maturation and IVM (P metabolism, apoptosis, extracellular matrix, and QTL-F-Fert-BTA3 genes were analyzed in surrounding cumulus cells (CC). In vivo, a significant decrease in prostaglandin synthase PTGES1 and PTGS2 expression coupled with lower PTGS2 protein abundance in CC and reduced expression of MOS in enclosed metaphase-II oocytes from "Fertil-" cows was observed. IVM strongly deregulated gene expression in CC and in oocytes compared with in vivo; nevertheless, differential expression of several genes including PEX19, NAMPT and MOS was observed between the two haplotypes. During IVM, PTGS2 activity inhibitor NS398 (50 μM) led to lower expression of fatty acid synthase (FASN) in CC and of MOS in treated metaphase-II oocytes. Using immunofluorescence, MOS protein was localized to a midbody-like contractile ring separating the polar body from the ooplasm, suggesting a role in the terminal stage of oocyte maturation. Our results suggest that factors involved in prostaglandin synthesis and lipid metabolism in CC could impair oocyte maturation, and might be involved in the reduced fertility of "Fertil-" cows. Copyright © 2014 Elsevier Inc. All rights reserved.

  17. Comparison of cytological image analysis systems in biological dosimetry

    International Nuclear Information System (INIS)

    Roy, L.; Delbos, M.; Paillole, N.; Durand, V.; Voisin, P.

    2003-01-01

    In biological dosimetry the reference technique is based on the scoring of dicentrics and centric rings induced by ionizing radiation. This article tends to present some image analysis systems used in biological dosimetry to help aberration detection. The presented systems are: the CYTOGEN from IMSTAR society, the CYTOSCAN (APPLIED IMAGING) and the METAFER (METASYSTEM). All 3 systems do not present similar functionalities but offer different way of automation. Some functionalities of these systems are compared. The systems can assist operators on 3 major points: (1) an automatic metaphase finder, in this case, the time benefic ranges from 2 to 4 when compared to manual scoring; (2) a specific tool to help manual scoring of aberrations, this provide an easier scoring and a better stability of the results; (3) the automatic detection of dicentrics, very useful in case of population triage. In this case, the dose has to be established very quickly, on a large population. However, the precision on the dose can be reduced compared to individual dose estimation. Some uncertainties on the detection of dicentric are acceptable and dicentric detection can be automated. Time benefic is then very important as 300 cells can be analyzed in half an hour (METAFER) against 25 when the metaphase finder is used alone. However, only 50% of the dicentrics are detected therefore it would be interesting to increase the efficiency of the detection. The use of the FISH technique could allow a better detection of dicentrics, the first results are promising (90% of centromeres correctly detected) but more experiments are required to evaluate the time benefit. (authors)

  18. The GnRH analogue triptorelin confers ovarian radio-protection to adult female rats

    International Nuclear Information System (INIS)

    Camats, N.; Garcia, F.; Parrilla, J.J.; Calaf, J.; Martin-Mateo, M.; Caldes, M. Garcia

    2009-01-01

    There is a controversy regarding the effects of the analogues of the gonadotrophin-releasing hormone (GnRH) in radiotherapy. This has led us to study the possible radio-protection of the ovarian function of a GnRH agonist analogue (GnRHa), triptorelin, in adult, female rats (Rattus norvegicus sp.). The effects of the X-irradiation on the oocytes of ovarian primordial follicles, with and without GnRHa treatment, were compared, directly in the female rats (F 0 ) with reproductive parameters, and in the somatic cells of the resulting foetuses (F 1 ) with cytogenetical parameters. In order to do this, the ovaries and uteri from 82 females were extracted for the reproductive analysis and 236 foetuses were obtained for cytogenetical analysis. The cytogenetical study was based on the data from 22,151 metaphases analysed. The cytogenetical parameters analysed to assess the existence of chromosomal instability were the number of aberrant metaphases (2234) and the number (2854) and type of structural chromosomal aberrations, including gaps and breaks. Concerning the reproductive analysis of the ovaries and the uteri, the parameters analysed were the number of corpora lutea, implantations, implantation losses and foetuses. Triptorelin confers radio-protection of the ovaries in front of chromosomal instability, which is different, with respect to the single and fractioned dose. The cytogenetical analysis shows a general decrease in most of the parameters of the triptorelin-treated groups, with respect to their controls, and some of these differences were considered to be statistically significant. The reproductive analysis indicates that there is also radio-protection by the agonist, although minor to the cytogenetical one. Only some of the analysed parameters show a statistically significant decrease in the triptorelin-treated groups.

  19. Drug-induced premature chromosome condensation (PCC) protocols: cytogenetic approaches in mitotic chromosome and interphase chromatin.

    Science.gov (United States)

    Gotoh, Eisuke

    2015-01-01

    Chromosome analysis is a fundamental technique which is used in wide areas of cytogenetic study including karyotyping species, hereditary diseases diagnosis, or chromosome biology study. Chromosomes are usually prepared from mitotic cells arrested by colcemid block protocol. However, obtaining mitotic chromosomes is often hampered under several circumstances. As a result, cytogenetic analysis will be sometimes difficult or even impossible in such cases. Premature chromosome condensation (PCC) (see Note 1) is an alternative method that has proved to be a unique and useful way in chromosome analysis. Former, PCC has been achieved following cell fusion method (cell-fusion PCC) mediated either by fusogenic viruses (e.g., Sendai virus) or cell fusion chemicals (e.g., polyethylene glycol), but the cell fusion PCC has several drawbacks. The novel drug-induced PCC using protein phosphatase inhibitors was introduced about 20 years ago. This method is much simpler and easier even than the conventional mitotic chromosome preparation protocol use with colcemid block and furthermore obtained PCC index (equivalent to mitotic index for metaphase chromosome) is usually much higher than colcemid block method. Moreover, this method allows the interphase chromatin to be condensed to visualize like mitotic chromosomes. Therefore drug-induced PCC has opened the way for chromosome analysis not only in metaphase chromosomes but also in interphase chromatin. The drug-induced PCC has thus proven the usefulness in cytogenetics and other cell biology fields. For this second edition version, updated modifications/changes are supplemented in Subheadings 2, 3, and 4, and a new section describing the application of PCC in chromosome science fields is added with citation of updated references.

  20. Analysis of chromosomal abnormalities: a study of partial exposure to X-rays

    Energy Technology Data Exchange (ETDEWEB)

    Andrade, Aida M.G. de; Mendes, Mariana E.; Mendonça, Julyanne C.G.; Melo, Laís; Hwang, Suy; Santos, Neide; Lima, Fabiana F. de, E-mail: aidamgandrade@gmail.com [Universidade Federal de Pernambuco (UFPE), Recife (Brazil); Centro Regional de Ciências Nucleares (CRCN-NE/CNEN-PE), Recife, PE (Brazil); Universidade Federal de Pernambuco (UFPE),Recife (Brazil). Centro de Biociências. Departamento de Genética

    2017-11-01

    Biological dosimetry is used in case of supposed accidental overexposure. The most commonly used biomarkers for assessing the absorbed dose are unstable chromosomal abnormalities. In a case of a partial body exposure, the frequencies of those abnormalities varies according to the area of the exposed body and may be substantially different from a total exposure of the body with an identical dose. The present study aimed to evaluate the frequency of chromosomal changes simulating, with blood samples, partial (25%, 50%) and full body irradiation (100%) in X-ray beam. The irradiation was performed at Metrology Service (CRCN-NE / CNEN) with a bundle of 250kVp X-rays, resulting in the absorbed dose of 1.0 Gy. Prior to obtain the metaphases, irradiated blood was mixed with non-irradiated blood, and then the mitotic metaphases for the chromosomal analyzes were obtained by culturing lymphocytes and the slides were stained with 5% Giemsa. It was observed that there was an increase in dicentric frequency when the dose percentage increases in both subjects (0.024 and 0.049 in subject 1 and 0.016 and 0.038 in subject 2) after irradiation. The cellular distribution was 'contaminated' only at dose 25% of the first individual who had a prolongation of the distribution. The Qdr and Dolphin methods were used to estimate partial absorbed dose, but the Qdr method was not efficient and whereas the Dolphin method was efficient when the individual had a prolonged cell distribution. It is necessary to increase the number of observations to be sure of the observed behaviors. (author)

  1. CYTOGENETIC STUDIES ON SEVERAL HIPPOPHAƠ RHAMNOIDES L. GENOTYPES

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    Diana-Elena Maftei

    2012-06-01

    Full Text Available Sea-buckthorn (Hippophaë rhamnoides (2n=24 is a dioecious plant with a very obvious morpho-physiological polymorphism. This species is one of the most valuable fruit-bearing shrubs of the spontaneous and also of the cultivated flora, due to its content of biologically active substances of its leaves, fruit and shoots. The research has been accomplished on root apical meristems from germinated seeds that belong to four genotypes characteristic for Bacău county - Dospineúti, ùerpeni 11, ùerbăneúti 4, Sfiútofca 18, and to one genotype of the Danube Delta - Sfântul Gheorghe 5. Cytogenetical studies evinced that the mitotic index (MI was high, and it varied with the genotype. The highest MI was evinced in the ùerbăneúti 4 genotype (53.13, and the lowest – in ùerpeni 11 (40.08. Cell distribution per mitotic phases is approximately the same, the highest percentage was represented by cells in prophase, followed by metaphases, telophases and anaphases. There is a rather high frequency and a quite large spectrum of  chromosomal aberrations identified in this species (with variations due to the 5 different genotypes. Of all the chromosomal aberrations evinced during the ana-telophases of mitotic root apical meristems (bridges, delayed chromosomes, expelled chromosomes, fragments, micronuclei, the highest frequency was represented by ana-telophases with bridges. There has been noticed the presence of chromosomal abnormalities in metaphasic and prophasic cells.

  2. Multiple requirements of PLK1 during mouse oocyte maturation.

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    Petr Solc

    Full Text Available Polo-like kinase 1 (PLK1 orchestrates multiple events of cell division. Although PLK1 function has been intensively studied in centriole-containing and rapidly cycling somatic cells, much less is known about its function in the meiotic divisions of mammalian oocytes, which arrest for a long period of time in prophase before meiotic resumption and lack centrioles for spindle assembly. Here, using specific small molecule inhibition combined with live mouse oocyte imaging, we comprehensively characterize meiotic PLK1's functions. We show that PLK1 becomes activated at meiotic resumption on microtubule organizing centers (MTOCs and later at kinetochores. PLK1 is required for efficient meiotic resumption by promoting nuclear envelope breakdown. PLK1 is also needed to recruit centrosomal proteins to acentriolar MTOCs to promote normal spindle formation, as well as for stable kinetochore-microtubule attachment. Consequently, PLK1 inhibition leads to metaphase I arrest with misaligned chromosomes activating the spindle assembly checkpoint (SAC. Unlike in mitosis, the metaphase I arrest is not bypassed by the inactivation of the SAC. We show that PLK1 is required for the full activation of the anaphase promoting complex/cyclosome (APC/C by promoting the degradation of the APC/C inhibitor EMI1 and is therefore essential for entry into anaphase I. Moreover, our data suggest that PLK1 is required for proper chromosome segregation and the maintenance of chromosome condensation during the meiosis I-II transition, independently of the APC/C. Thus, our results define the meiotic roles of PLK1 in oocytes and reveal interesting differential requirements of PLK1 between mitosis and oocyte meiosis in mammals.

  3. Sesamol attenuates cytogenetic damages in bone marrow cells of whole body gamma irradiated mice

    International Nuclear Information System (INIS)

    Kumar, Arun; Tamizh Selvan, G.; Adhikari, Jawahar S.; Chaudhury, N.K.

    2014-01-01

    Whole body radiation exposure cause damages to all vital organs and bone marrow is the most sensitive. Pre-treatment with antioxidant as single prophylactic dose is expected to lower induction of damages in bone marrow. In the present study we have focused on sesamol, a dietary antioxidant mediated radioprotection in bone marrow cells of gamma irradiated mice and compared with melatonin. Male C57BL/6 mice were intraperitoneally administered with sesamol (10 and 20 mg/kg body) and after 30 minutes exposed to whole body gamma radiation using 60 Co Teletherapy unit. Mice were injected with 0.2 ml of a metaphase arresting agent (0.05% colchicine) intra-peritoneally 3 hours prior to sacrifice (24 hrs. post-irradiation). Bone marrow cells were flushed out from femurs of each animal and processed for chromosomal aberration assay. Another set of experiment without colchicine injection was performed to access the DNA damage in bone marrow using alkaline comet assay. At least 100 metaphases per animal were scored under light microscope to record various aberrations and total chromosomal aberrations (TCA) was calculated. Similar measurements were performed with melatonin for comparing the efficacy of sesamol. Gamma irradiation has increased the chromatid type aberrations (break formation, fragment) and chromosomal type aberrations (ring formation, acentric) in bone marrow cells. The results have shown significant (p< 0.001) increase in TCA of irradiated mice than control. While pre-treatment of sesamol and melatonin 10 mg/kg significantly (p<0.05) reduced the TCA. The extend of protection has increased at 20 mg/kg significantly (p<0.001) as evident from the reduced TCA compared to irradiated group. Interestingly, sesamol and melatonin have shown similar extent of reduction of TCA. Thus sesamol has demonstrated strong ability to protect bone marrow at low dosage. These investigations on sesamol mediated protection in bone marrow are likely to benefit development of

  4. Role of cytogenetic biodosimetry in meeting the needs of a mass casualty radiological/nuclear event

    International Nuclear Information System (INIS)

    Balajee, A.S.; Dainiak, N.

    2016-01-01

    Radiological/nuclear (R/N) terrorism constitutes a potential threat to all nations that can result in significant morbidity and mortality among hundreds of thousands individuals. In addition to the timing and severity of clinical signs and symptoms, individual radiation dose informs risk assessment and mitigation of radiation-associated injuries. The 'gold standard' for individual whole-body radiation dosimetry is the dicentric chromosome assay. The Cytogenetics Biodosimetry Laboratory at REAC/TS is a WHO Collaborating Centre and member of IAEA's RANET that employs DCA, as well as fluorescence in situ hybridization, premature chromosome condensation, and micronuclei assays to assess radiation dose. The quality of dose estimates and standard operating procedures for DCA at REAC/TS have been validated in multiple inter-comparison studies involving CBLs in Asia, Europe, North America and South America. DCA is scalable to meet the needs of a mass casualty R/N incident. The CBL at REAC/TS has made seminal contributions to augment surge capacity for DCA and develop CBLs worldwide through initiatives such as modification of 'Share Point' in 2010 to transmit images of metaphases for simultaneous telescoring; (2) development of an on-line training program for metaphase scoring; (3) proactive participation as a WCC to create ISO standards; and (4) guidance of regulatory agencies to monitor quality of results and SOPs. The precision of dose estimates by DCA can be vastly improved by using a universal calibration curve. With this view, REAC/TS has organized a collaboration with CBLs at Health Canada and Yale University to construct and validate a common calibration curve for gamma rays

  5. Identification of a TPX2-like microtubule-associated protein in Drosophila.

    Directory of Open Access Journals (Sweden)

    Gohta Goshima

    Full Text Available Chromosome segregation during mitosis and meiosis relies on the spindle and the functions of numerous microtubule-associated proteins (MAPs. One of the best-studied spindle MAPs is the highly conserved TPX2, which has been reported to have characteristic intracellular dynamics and molecular activities, such as nuclear localisation in interphase, poleward movement in the metaphase spindle, microtubule nucleation, microtubule stabilisation, microtubule bundling, Aurora A kinase activation, kinesin-5 binding, and kinesin-12 recruitment. This protein has been shown to be essential for spindle formation in every cell type analysed so far. However, as yet, TPX2 homologues have not been found in the Drosophila genome. In this study, I found that the Drosophila protein Ssp1/Mei-38 has significant homology to TPX2. Sequence conservation was limited to the putative spindle microtubule-associated region of TPX2, and intriguingly, D-TPX2 (Ssp1/Mei-38 lacks Aurora A- and kinesin-5-binding domains, which are highly conserved in other animal and plant species, including many insects such as ants and bees. D-TPX2 uniformly localised to kinetochore microtubule-enriched regions of the metaphase spindle in the S2 cell line, and it had microtubule binding and bundling activities in vitro. In comparison with other systems, the contribution of D-TPX2 to cell division seems to be minor; live cell imaging of microtubules and chromosomes after RNAi knockdown identified significant delay in chromosome congression in only 18% of the cells. Thus, while this conserved spindle protein is present in Drosophila, other mechanisms may largely compensate for its spindle assembly and chromosome segregation functions.

  6. Casein kinase 1 alpha regulates chromosome congression and separation during mouse oocyte meiotic maturation and early embryo development.

    Directory of Open Access Journals (Sweden)

    Lu Wang

    Full Text Available Casein kinase I alpha (CK1α is a member of serine/threonine protein kinase, generally present in all eukaryotes. In mammals, CK1α regulates the transition from interphase to metaphase in mitosis. However, little is known about its role in meiosis. Here we examined Ck1α mRNA and protein expression, as well as its subcellular localization in mouse oocytes from germinal vesicle to the late 1-cell stage. Our results showed that the expression level of CK1α was increased in metaphase. Immunostaining results showed that CK1α colocalized with condensed chromosomes during oocyte meiotic maturation and early embryo development. We used the loss-of-function approach by employing CK1α specific morpholino injection to block the function of CK1α. This functional blocking leads to failure of polar body 1 (PB1 extrusion, chromosome misalignment and MII plate incrassation. We further found that D4476, a specific and efficient CK1 inhibitor, decreased the rate of PB1 extrusion. Moreover, D4476 resulted in giant polar body extrusion, oocyte pro-MI arrest, chromosome congression failure and impairment of embryo developmental potential. In addition, we employed pyrvinium pamoate (PP, an allosteric activator of CK1α, to enhance CK1α activity in oocytes. Supplementation of PP induced oocyte meiotic maturation failure, severe congression abnormalities and misalignment of chromosomes. Taken together, our study for the first time demonstrates that CK1α is required for chromosome alignment and segregation during oocyte meiotic maturation and early embryo development.

  7. Dynamics of tobacco DNA topoisomerases II in cell cycle regulation: to manage topological constrains during replication, transcription and mitotic chromosome condensation and segregation.

    Science.gov (United States)

    Singh, Badri Nath; Achary, V Mohan Murali; Panditi, Varakumar; Sopory, Sudhir K; Reddy, Malireddy K

    2017-08-01

    The topoisomerase II expression varies as a function of cell proliferation. Maximal topoisomerase II expression was tightly coupled to S phase and G2/M phase via both transcriptional and post-transcriptional regulation. Investigation in meiosis using pollen mother cells also revealed that it is not the major component of meiotic chromosomes, it seems to diffuse out once meiotic chromosomal condensation is completed. Synchronized tobacco BY-2 cell cultures were used to study the role of topoisomerase II in various stages of the cell cycle. Topoisomerase II transcript accumulation was observed during the S- and G2/M- phase of cell cycle. This biphasic expression pattern indicates the active requirement of topoisomerase II during these stages of the cell cycle. Through immuno-localization of topoisomerase II was observed diffusely throughout the nucleoplasm in interphase nuclei, whereas, the nucleolus region exhibited a more prominent immuno-positive staining that correlated with rRNA transcription, as shown by propidium iodide staining and BrUTP incorporation. The immuno-staining analysis also showed that topoisomerase II is the major component of mitotic chromosomes and remain attached to the chromosomes during cell division. The inhibition of topoisomerase II activity using specific inhibitors revealed quite dramatic effect on condensation of chromatin and chromosome individualization from prophase to metaphase transition. Partially condensed chromosomes were not arranged on metaphase plate and chromosomal perturbations were observed when advance to anaphase, suggesting the importance of topoisomerase II activity for proper chromosome condensation and segregation during mitosis. Contrary, topoisomerase II is not the major component of meiotic chromosomes, even though mitosis and meiosis share many processes, including the DNA replication, chromosome condensation and precisely regulated partitioning of chromosomes into daughter cells. Even if topoisomerase II is

  8. Chromosome stickiness during meiotic behavior analysis of Passiflora serrato-digitata L. (PassifloraCEAE Aderência cromossômica durante a análise do comportamento meiótico de Passiflora serrato-digitata L (PassifloraCEAE

    Directory of Open Access Journals (Sweden)

    Paulo Roberto Peres Kiihl

    2011-06-01

    Full Text Available Almost 90% of species of the genus Passiflora are native to the American continent, with high commercial value due to the fact that some species are used for human food while others have ornamental and medical qualities. Passiflora serrato-digitata is one of the species that integrates the Paraná Agronomic Institute germoplasm bank at its experimental base in Londrina, PR, Brazil. Collected flower buds were fixed in ethanol/acetic acid (3:1 v/v for 24h, transferred to 70% alcohol and stored under refrigeration. Slides were prepared by the squashing technique and stained with 1.0% propionic carmine; they were analyzed under an optic microscope. Irregularities in the chromosome segregation process of P. serrato-digitata have been verified by meiotic behavior analysis. These comprised precocious migration to poles in metaphase I and II, non-oriented chromosomes in metaphase plate in metaphase I and II, laggard chromosomes in anaphase I and II towards the formation of micronucleus in telophase I and II, and microspores in tetrads. Chromosome stickiness was another irregularity reported in the Passiflora genus for the first time. These irregularities which also contributed to the formation of monads, dyads and triads, resulted in normal imbalanced 2n and 4n microspores. According to the observed Meiotic Index of 71.83%, this species is not meiotically stable.Cerca de 90% das espécies do gênero Passiflora são nativas das Américas, sendo que aproximadamente 200 espécies são nativas do Brasil. Possuem grande importância comercial, pois algumas espécies são utilizadas na alimentação humana, outras apresentam propriedades medicinais e ornamentais. A espécie Passiflora serrato-digitata faz parte do banco de germoplasma do Instituto Agronômico do Paraná - IAPAR, estação experimental de Londrina, PR. Botões florais colhidos foram fixados em etanol/ácido acético (3:1 v/v por 24 horas, transferidos para álcool a 70% e acondicionado sob

  9. Classic and molecular cytogenetic analysis regarding human reactivity to beta radiation

    International Nuclear Information System (INIS)

    Usurelu Daniela; Radu Irina; Gavrila Lucian; Cimponeriu Danut; Apostol Pompilia; Ahmadi Elham

    2007-01-01

    , dicentric chromosomes, monochromatidic gap and so on. The most important targets for ionizing radiation being the telomeres, we've tested from this point of view the integrity/modification of telomeres by FISH technique. After we analyzed 100 fluorescent metaphases/irradiation doses/normal case we found the fluorescent signal in approx. 98% of the normal metaphases. After the irradiation, in cases like complex chromosomal interchange or telomere to telomere translocation we did not identified the fluorescent signal at the chromosomal ends implicated in the rearrangements. This observation and the fact that we found the signal on different acentric fragments revealed that the beta irradiation generates important destruction at the chromosomal end. In this study we've also analyzed one of the telomerase compounds - RNA compound, in normal condition and after beta irradiation by FISH technique. After the molecular cytogenetic analysis we identified the RNA telomerase compound on chromosome 3, q arm, at the two homologue chromosomes in normal probes and also in irradiated one. The fact that we did not found additional signals (more than four signal/metaphase) after irradiation with beta rays revealed the fact, that in this case, the telomerase is not amplified in order to repair the broken telomeres.

  10. Differences and homologies of chromosomal alterations within and between breast cancer cell lines: a clustering analysis.

    Science.gov (United States)

    Rondón-Lagos, Milena; Verdun Di Cantogno, Ludovica; Marchiò, Caterina; Rangel, Nelson; Payan-Gomez, Cesar; Gugliotta, Patrizia; Botta, Cristina; Bussolati, Gianni; Ramírez-Clavijo, Sandra R; Pasini, Barbara; Sapino, Anna

    2014-01-23

    The MCF7 (ER+/HER2-), T47D (ER+/HER2-), BT474 (ER+/HER2+) and SKBR3 (ER-/HER2+) breast cancer cell lines are widely used in breast cancer research as paradigms of the luminal and HER2 phenotypes. Although they have been subjected to cytogenetic analysis, their chromosomal abnormalities have not been carefully characterized, and their differential cytogenetic profiles have not yet been established. In addition, techniques such as comparative genomic hybridization (CGH), microarray-based CGH and multiplex ligation-dependent probe amplification (MLPA) have described specific regions of gains, losses and amplifications of these cell lines; however, these techniques cannot detect balanced chromosomal rearrangements (e.g., translocations or inversions) or low frequency mosaicism. A range of 19 to 26 metaphases of the MCF7, T47D, BT474 and SKBR3 cell lines was studied using conventional (G-banding) and molecular cytogenetic techniques (multi-color fluorescence in situ hybridization, M-FISH). We detected previously unreported chromosomal changes and determined the content and frequency of chromosomal markers. MCF7 and T47D (ER+/HER2-) cells showed a less complex chromosomal make up, with more numerical than structural alterations, compared to BT474 and SKBR3 (HER2+) cells, which harbored the highest frequency of numerical and structural aberrations. Karyotype heterogeneity and clonality were determined by comparing all metaphases within and between the four cell lines by hierarchical clustering. The latter analysis identified five main clusters. One of these clusters was characterized by numerical chromosomal abnormalities common to all cell lines, and the other four clusters encompassed cell-specific chromosomal abnormalities. T47D and BT474 cells shared the most chromosomal abnormalities, some of which were shared with SKBR3 cells. MCF7 cells showed a chromosomal pattern that was markedly different from those of the other cell lines. Our study provides a comprehensive

  11. lemmingA encodes the Apc11 subunit of the APC/C in Drosophila melanogaster that forms a ternary complex with the E2-C type ubiquitin conjugating enzyme, Vihar and Morula/Apc2

    Directory of Open Access Journals (Sweden)

    Nagy Olga

    2012-03-01

    Full Text Available Abstract Background Ubiquitin-dependent protein degradation is a critical step in key cell cycle events, such as metaphase-anaphase transition and mitotic exit. The anaphase promoting complex/cyclosome (APC/C plays a pivotal role in these transitions by recognizing and marking regulatory proteins for proteasomal degradation. Its overall structure and function has been elucidated mostly in yeasts and mammalian cell lines. The APC/C is, however, a multisubunit assembly with at least 13 subunits and their function and interaction within the complex is still relatively uncharacterized, particularly in metazoan systems. Here, lemming (lmg mutants were used to study the APC/C subunit, Apc11, and its interaction partners in Drosophila melanogaster. Results The lmg gene was initially identified through a pharate adult lethal P element insertion mutation expressing developmental abnormalities and widespread apoptosis in larval imaginal discs and pupal abdominal histoblasts. Larval neuroblasts were observed to arrest mitosis in a metaphase-like state with highly condensed, scattered chromosomes and frequent polyploidy. These neuroblasts contain high levels of both cyclin A and cyclin B. The lmg gene was cloned by virtue of the lmg03424 P element insertion which is located in the 5' untranslated region. The lemming locus is transcribed to give a 2.0 kb mRNA that contains two ORFs, lmgA and lmgB. The lmgA ORF codes for a putative protein with more than 80% sequence homology to the APC11 subunit of the human APC/C. The 85 amino acid protein also contains a RING-finger motif characteristic of known APC11 subunits. The lmgA ORF alone was sufficient to rescue the lethal and mitotic phenotypes of the lmg138 null allele and to complement the temperature sensitive lethal phenotype of the APC11-myc9 budding yeast mutant. The LmgA protein interacts with Mr/Apc2, and they together form a binding site for Vihar, the E2-C type ubiquitin conjugating enzyme. Despite

  12. Utilização da contagem de folículos antrais para predição do padrão de resposta em ciclos de hiperestimulação controlada com antagonista de GnRH Use of antral follicle count to predict the response pattern in controlled ovarian hyperstimulation cycles with GnRH antagonist

    Directory of Open Access Journals (Sweden)

    Maria do Carmo Borges de Souza

    2008-01-01

    Full Text Available OBJETIVO: verificar se existe relação preditiva entre a contagem de folículos antrais (CFA no segundo dia do ciclo com o padrão de resposta em ciclos de hiperestimulação ovariana controlada para injeção intracitoplasmática de espermatozóide (ICSI. MÉTODOS: estudo prospectivo, desenvolvido de maio de 2004 a maio de 2005, no qual 51 pacientes com idade 15 mm no dia do desencadeamento da ovulação, número total e em metáfase II de oócitos captados, número de embriões de boa qualidade transferidos e taxa de gestação. A análise estatística foi realizada pelos testes t de Student e de Mann-Whitney, com significância estatística de 5% (pPURPOSE: to establish whether there is a predictive relationship between the antral follicle count (AFC on the second day of the cycle and the response pattern in controlled ovarian hyperstimulation cycles for intracytoplasmic sperm injection (ICSI. METHODS: a prospective study developed from May 2004 to May 2005, in which 51 patients aged 15 mm on the day of ovulation triggering, the total number of oocytes retrieved and in metaphases II, the number of good quality embryos transferred and pregnancy rate. The statistical analysis was performed by the t-Student test and the Mann-Whitney test, with statistical significance of 5% (p15 mm on the day of ovulation triggering (p=0.0001, the total number of oocytes retrieved (p=0.0001 and those in metaphases II (p=0.0001. Such correlation between AFC and pregnancy was not observed (p=0.43. There was no significant correlation between AFC and the number of good quality embryos transferred (p=0.081. CONCLUSIONS: AFC on the second day of the stimulated cycle can be used to predict the quality of ovarian stimulation, the number of oocytes retrieved and the number of mature oocytes in in vitro fertilization cycles using GnRH antagonist.

  13. Potent radio-protective effects of vitamins E and C on radiation induced DNA damage in gametes leading to lower frequencies of chromosomal aberrations and micronuclei in subsequent embryos

    International Nuclear Information System (INIS)

    Hossein Mozdarani

    2007-01-01

    Complete text of publication follows. Objective: To compare the effects of parental and maternal exposure of NMRI mice with γ-rays on gametes in the absence or presence of vitamins E and C and subsequent cytogenetic damage in pre-implantation embryos generated from irradiated gametes. Materials and Methods: Male and female NMRI mice were whole body irradiated in the presence of 200 IU/Kg vitamin E and 100 μg/ml vitamin C. Various mating schemes were designed for mating of irradiated mice, e.g. mating irradiated male with non-irradiated female, irradiated female with non irradiated male or both male and female irradiated. About 68 h post coitus, 4-8-cell embryos were flushed out from oviducts and fixed on slides using standard methods in order to screen for chromosome abnormalities and micronuclei. Results: In control embryos, frequencies of abnormal metaphase and embryos with micronuclei was low and there was no significant difference between vitamins treated samples and controls. However there was an increase in both abnormal metaphases and micronuclei frequency in embryos generated after parental or maternal irradiation or both. Vitamin E effectively reduced the frequency of aneuploidy in all irradiated groups and vitamin C was very effective in reducing the frequencies of micronuclei. DRF calculated for both vitamins indicate that vitamin C is more potent than vitamin E in reducing clastogenic effects of gamma-rays in pre-implantation embryos. Conclusion: Data indicate that γ-irradiation affects spermatogenesis and preovulatory stage oocytes in male and female mice respectively. These effects might be due to DNA alterations in sperms and oocytes affecting meiotic segregations that may lead to chromosome abnormalities in subsequent embryos expressed as numerical chromosome abnormalities or micronuclei. Administration of vitamins E and C before irradiation effectively reduced the frequency of chromosomal abnormalities. The way these vitamins reduces genotoxic

  14. Cytogenetic aspects of a canine breast carcinosarcoma - a case report.

    Science.gov (United States)

    Morais, C S D; Affonso, P R A M; Bitencourt, J A; Wenceslau, A A

    2017-05-25

    This study searched a rare and aggressive type of cancer in dogs and humans, the breast carcinosarcoma. Both clinical and pathological traits of mammary carcinosarcomas in dogs are similar to humans, such as infrequent occurrence, fast tumor growth, and unfavorable prognosis when compared to carcinomas. Other possible alterations include chromosomal abnormalities that can be useful for the identification of tumoral cells and diagnosis. The aim of this study was to compare the chromosomal features of peripheral lymphocytes and tumor cells in a mammary carcinosarcoma of a 14-year-old female Poodle. Chromosomes were analyzed from 210 metaphases by conventional Giemsa staining, C-banding, and base-specific fluorochrome staining with chromomycin A3 (CMA 3 + ) and DAPI. Of the 105 blood cells, 56.3% followed the standard karyotype of dogs (2n = 78). In contrast, the carcinosarcoma cells showed high chromosomal numbers (104 to 153), divided into 80% hypertriploid (118 to 136 chromosomes), 10.5% hypotetraploid (137 to 153 chromosomes), 5.7% hypotriploid (104 to 116 chromosomes), and 3.8% triploid cells (117 chromosomes). Among the aneuploid cells identified, we highlighted the trisomy of pair 1 and X chromosome once these elements were easily recognized in karyotype because of their size (pair 1) or differential morphology. Heterochromatin in normal cells was restricted to the pericentromeric region of all chromosomes while few C-bands were observed in tumor cells. This apparent loss of heterochromatin in neoplastic cells was supposed to favor centric fusion among formerly acrocentric chromosomes. Fluorochrome staining reinforced this hypothesis once GC-rich segments (CMA 3 + ) were identified on 10 chromosomes from normal cells (2n = 78) whereas carcinosarcoma metaphases had up to 11 chromosomes bearing CMA 3 signals in spite of their remarkable high chromosomal numbers. We concluded that, like in humans, the carcinosarcoma in dogs caused genome instability that

  15. Nicotine-induced Disturbances of Meiotic Maturation in Cultured Mouse Oocytes: Alterations of Spindle Integrity and Chromosome Alignment

    Directory of Open Access Journals (Sweden)

    Zenzes Maria

    2004-09-01

    Full Text Available Abstract We investigated whether nicotine exposure in vitro of mouse oocytes affects spindle and chromosome function during meiotic maturation (M-I and M-II. Oocytes in germinal vesicle (GV stage were cultured in nicotine for 8 h or for 16 h, to assess effects in M-I and in metaphase II (M-II. The latter culture setting used the three protocols: 8 h nicotine then 8 h medium (8N + 8M; 16 h nicotine (16N; 8 h medium then 8 h nicotine (8M + 8N. Non-toxic concentrations of nicotine at 1.0, 2.5, 5.0 and 10.0 mmol/L were used. Spindle-chromosome configurations were analyzed with wide-field optical sectioning microscopy. In 8 h cultures, nicotine exposure resulted in dose-related increased proportions of M-I oocytes with defective spindle-chromosome configurations. A dose-related delayed entry into anaphase I was also detected. In 16 h cultures, nicotine exposure for the first 8 h (8N + 8M, or for 16 h (16N, resulted in dose- and time-related increased proportions of oocytes arrested in M-I (10 mmol/L; 8 h: 53.2%, controls 9.6%; 16 h: 87.6%, controls 8.5%. Defects in M-I spindles and chromosomes caused M-I arrest leading to dose-related decreased proportions of oocytes that reached metaphase-II (10 mmol/L 8 h: 46.8%, controls 90.4%;16 h: 12.4%, controls 91.5%. A delayed anaphase-I affected the normal timing of M-II, leading to abnormal oocytes with dispersed chromosomes, or with double spindles and no polar body. Nicotine exposure during the second 8 h (8M + 8N resulted in dose-related, increased proportions of M-II oocytes with defective spindles and chromosomes (10 mmol/L: 42.9%, controls 2.0%. Nicotine has no adverse effects on GV break down, but induces spindle and chromosome defects compromising oocyte meiotic maturation and development.

  16. Evaluation of chromosome aberration frequency instable in individual groups residents at the municipality of Monte Alegre, Para, Brazil, exposed to radon

    International Nuclear Information System (INIS)

    Yunes, Samira Nogarol

    2010-01-01

    The municipality of Monte Alegre is a region that presents natural radiation high due to the presence of the radionuclide uranium ( 238 U) in its soil, which through its decay gives rise to element Rn, a gas. The radioactivity of the rocks has become a problem for the population of Monte Alegre, from the moment when the radioactive material began to be used in the construction of houses and paving of streets. Among all bio markers related to environmental exposures and its biological effects, the chromosomal aberrations are considered good bio markers as predictors of the risk of cancer. Studies suggest that the frequency of chromosomal aberrations may be related to the genetic instability individual and/or exposure to ionizing radiation. Our work aimed to evaluate the frequency of chromosomal aberrations in individuals in the region of high natural radioactivity in Monte Alegre-PA. As well as to correlate the cytogenetic analysis made in this study with the results of analysis of frequency of polymorphisms of genes of DNA repair carried out in another study that resulted in other dissertation. In accordance with the distribution of the data obtained in characterizing environmental radiological and in the calculation of dose, were chosen residents of homes with more and less exposure to radiation. The samples of peripheral blood of 85 individuals of the resident population of the region of Monte Alegre - PA were collected and examine provided two slides for individual was performed to verify the quality of the sample. Through this evaluation we decide that 33% of the material collected, or is, samples of 28 individuals were in suitable conditions for analysis of the frequency of chromosomal aberrations. After the collections lymphocytes present in the sample were cultivated in accordance with the methodology proposed for obtaining of cells in metaphase. were analyzed 6,177 metaphases of 28 individuals among which were found dicentric chromosomes 4 and 19 fragments

  17. PRENATAL DIAGNOSIS AND SCREENING OF GENETIC ABNORMALITIES IN EARLY PREGNANCY

    Directory of Open Access Journals (Sweden)

    Jyothi Kiran Kohli

    2016-11-01

    Full Text Available BACKGROUND Genetic diseases are one of the major causes of hospital admissions due to disability and mortality particularly among children (1:5 children of hospital admission either partially/completely as distribution of genetic diseases is not related to socioeconomic background, which implies that developing world has a large number of genetic diseases largely left uncared for, i.e. overall incidence of foetal/neonatal loss due to genetic/genetic environmental causes are as follows: 1:50 newborns have major congenital abnormality, 1:100 have a unifactorial disorder, 1:200 have a major chromosomal abnormality before birth. Diagnosis of chromosomal anomalies in foetus is one of the most important challenges in modern perinatology as invasive or noninvasive methods. The aim of the study is to review on cytogenetic evaluation of CVS obtained (transcervically during first trimester of pregnancy by direct karyotyping of tissue. MATERIALS AND METHODS This study was conducted in 2001 in Department of Anatomy along with Obstetrics and Gynaecology Department, LNJP Hospital. 37 healthy cases with 6-12 weeks of gestational age coming for medical termination of pregnancy were included in the study. After written informed consent for procedure, ultrasound-guided transcervical chorionic villus sampling was done (Brambati’s method. Tissue procured was then processed for direct karyotyping and studied. Metaphase spreads were photographed and karyotypes prepared and studied. RESULTS Out of 37 pregnant females, 30 samples were successfully prepared and processed by Direct method out of which 23 were normal female (46, XX and 7 were normal male (46, XY. No normal anomaly was detected. Best biopsies were obtained with 8-12 weeks gestation. G Banding could not be performed as chromosome obtained were found to be resistant to banding. CONCLUSIONS To summarise chromosome preparations obtained from CVS by Direct method has advantage of providing sufficient number

  18. Chromatin compaction by condensin I, intra-kinetochore stretch and tension, and anaphase onset, in collective spindle assembly checkpoint interaction

    International Nuclear Information System (INIS)

    Matsson, Leif

    2014-01-01

    The control mechanism in mitosis and meiosis by which cells decide to inhibit or allow segregation, the so-called spindle assembly checkpoint (SAC), increases the fidelity of chromosome segregation. It acts like a clockwork mechanism which measures time in units of stable attachments of microtubules (MTs) to kinetochores (the order parameter). Stable MT–kinetochore attachments mediate poleward forces and ‘unstable’ attachments, acting alone or together with motor proteins on kinetochores via chromosomes, antipoleward forces. Stable and unstable attachments could be separated, and the non-equilibrium integrated MT mediated force acting on stably attached kinetochores was derived in a collective interaction (Matsson 2009 J. Phys.: Condens. Matter 21 502101), in which kinetochores were treated as rigid protein complexes. As forces and tension in that model became equally distributed in all bioriented sister chromatid (SC) pairs, segregation was inhibited without need of a ‘wait-anaphase’ signal. In this generalization, the kinetochore is divided into an inner chromatin proximal complex and an outer MT proximal complex, and the integrated MT mediated force is divided into an integrated poleward and an integrated antipoleward force. The model also describes the collective interaction of condensin I with chromatin, which together with the MT mediated dynamics yields the putative in vivo tension in kinetochores and centromeric and pericentromeric chromatin, as a non-linear function of the order parameter. Supported by the compaction force and an increased stiffness in chromatin towards the end of metaphase, the two opposing integrated MT mediated poleward forces, together with metaphase oscillations, induce a swift and synchronized anaphase onset by first increasing the intra-kinetochore stretch. This increase lowers the SAC energy threshold, making a cleavage by separase of all cohesin tethering SC pairs in anaphase energetically possible, thereby reducing the

  19. Function of donor cell centrosome in intraspecies and interspecies nuclear transfer embryos

    International Nuclear Information System (INIS)

    Zhong Zhisheng; Zhang Gang; Meng Xiaoqian; Zhang Yanling; Chen Dayuan; Schatten, Heide; Sun Qingyuan

    2005-01-01

    Centrosomes, the main microtubule-organizing centers (MTOCs) in most animal cells, are important for many cellular activities such as assembly of the mitotic spindle, establishment of cell polarity, and cell movement. In nuclear transfer (NT), MTOCs that are located at the poles of the meiotic spindle are removed from the recipient oocyte, while the centrosome of the donor cell is introduced. We used mouse MII oocytes as recipients, mouse fibroblasts, rat fibroblasts, or pig granulosa cells as donor cells to construct intraspecies and interspecies nuclear transfer embryos in order to observe centrosome dynamics and functions. Three antibodies against centrin, γ-tubulin, and NuMA, respectively, were used to stain the centrosome. Centrin was not detected either at the poles of transient spindles or at the poles of first mitotic spindles. γ-tubulin translocated into the two poles of the transient spindles, while no accumulated γ-tubulin aggregates were detected in the area adjacent to the two pseudo-pronuclei. At first mitotic metaphase, γ-tubulin was translocated to the spindle poles. The distribution of γ-tubulin was similar in mouse intraspecies and rat-mouse interspecies embryos. The NuMA antibody that we used can recognize porcine but not murine NuMA protein, so it was used to trace the NuMA protein of donor cell in reconstructed embryos. In the pig-mouse interspecies reconstructed embryos, NuMA concentrated between the disarrayed chromosomes soon after activation and translocated to the transient spindle poles. NuMA then immigrated into pseudo-pronuclei. After pseudo-pronuclear envelope breakdown, NuMA was located between the chromosomes and then translocated to the spindle poles of first mitotic metaphase. γ-tubulin antibody microinjection resulted in spindle disorganization and retardation of the first cell division. NuMA antibody microinjection also resulted in spindle disorganization. Our findings indicate that (1) the donor cell centrosome, defined as

  20. Comparative study of dose-response curve for chromosome aberrations induced in human lymphocytes by {sup 60}Co and X-Rays

    Energy Technology Data Exchange (ETDEWEB)

    Mendes, Mariana E.; Mendonça, Julyanne C.G.; Andrade, Aida M.G.; Silva, Laís M.; Hwang, Suy; Melo, Ana M.M.A.; Santos, Neide; Lima, Fabiana F., E-mail: mendes_sb@hotmail.com [Centro Regional de Ciencias Nucleares (CRCN-NE/CNEN-PE), Recife, PE (Brazil); Universidade Federal de Pernambuco (UFPE), Recife, PE (Brazil)

    2017-11-01

    Biodosimetry represents a biological marker to the estimation of health risks after accidental overexposure to ionizing radiation. Chromosomal dicentric in peripheral blood lymphocytes have been the most reliable biomarker of exposure to IR during the last several decades. This technique could be used to support physical dosimetry or when it is impossible to achieve it. A reliable measurement of the absorbed dose is critical for medical decision, including the assessment of long-term health consequences. The aim of this research is to compare dose-response curves for dicentric aberration induced in human lymphocytes by {sup 60}Co and X-Rays. For both quality of radiation, the samples were exposed to at least eight different absorbed doses. The X-rays with dose rate of 0,275 Gy/min at Laboratory of Metrology (CRCN/NE - PE - Brazil) and the second one was exposed to cobalt source with dose rate of 0.055 Gy/min ({sup 60}Co Gammacell 220) located at Department of Nuclear Energy (UFPE-DEN-BRASIL). Mitotic metaphase cells were obtained by lymphocyte culture for chromosomal analysis and slides were stained with Giemsa 5%. The frequencies of dicentrics were counted in more than 18.000 metaphases for this comparison. After that, all frequencies and distributions of dicentrics were tested to analyze their conformity with Poisson distribution and then each quality of radiation were used for build the calibration curves using Dose Estimate program. These results showed that both curves followed the Poisson distribution and coefficients of each one are: YX-rays = 0,0013 (± 0,0006) + 0,0271 (± 0,0086)⁎D + 0,0556(±0,0050))⁎D{sup 2} and Y{sub Co-60} = 0,0014 (± 0,0010) + 0,0081 (± 0,0073))⁎D + 0,0451 (± 0,0046))⁎D{sup 2} (Y = frequency of dicentrics and D = absorbed dose). It was expected that there was no significant difference between this two types of radiation because both were low LET. We believed that dose rate have been a principal factor to produce this