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PubMed1991

Chromosome analysis of multipronuclear human oocytes after in vitro fertilization.

Multipronuclear human eggs are frequent after in vitro fertilization. Their chromosome analysis can provide useful information. Before cleavage it can confirm the suspected polyploidy. Among the cleaved multipronuclear eggs it provides an estimation of the incidence of the possible return to diploidy. Ninety-four multipronuclear eggs were fixed at the first, second, or third cleavage according to the air-drying method of Tarkowski with or without colchicine exposure: 60 were successfully analysed. Twelve were stopped before cleavage (six without colchicine treatment and six with colchicine treatment). They were polyploid, confirming the cytological observation. Forty-eight eggs cleaved and were stopped by colchicine treatment and karyotyped. Seventeen eggs (35 per cent) had produced diploid embryos. Mosaicism was frequent (15 cases, 31 per cent). Triploidy was not frequent (8 eggs, 17 per cent). Haploidy constituted the remaining cases (8 eggs, 17 per cent). Our data indicate that the initial count of pronuclei is a reliable test. Multipronuclear one-cell oocytes were confirmed to be polyploid. Furthermore, the developmental capacity of the multipronuclear oocytes is variable. Most of them cleaved. However, many multipronuclear oocytes led to diploid cleaving eggs.

Adult↗
PubMed1993

Cytogenetic analysis of human oocytes after subzonal insemination.

Subzonal insemination has been proposed to achieve fertilization in cases where standard in vitro fertilization has failed. We present the results of chromosome analysis of oocytes after subzonal insemination. Our data suggest that the main cause (76 per cent) of the absence of cleavage after subzonal insemination is the total absence of sperm nucleus evolution of the injected spermatozoa. Our results also suggest that spermatozoa chromatin development is normal after subzonal insemination. Aneuploidy does not seem to be increased in zygotes after subzonal insemination. However, polyploidy was often more important than predicted by the observation of pronuclei (PN). Pronucleus development might be asynchronous and can appear earlier or later than after standard IVF. The cytogenetic risk after subzonal insemination might therefore be triploidy (if a triploid egg is transferred, because only 2 PN were seen) rather than aneuploidy or structural abnormalities.

Cell Nucleus↗
PubMed1996

Overexpression of granulocyte colony-stimulating factor in vivo decreases the level of polyploidization of mouse bone marrow megakaryocytes.

The in vivo effect of G-CSF on the maturation of mouse bone marrow megakaryocytes was studied by monitoring the DNA contents. Megakaryocytes were first identified by a specific 1C2 monoclonal antibody against mouse platelets and megakaryocytes and DNA contents of these cells were measured by propidium iodine. Megakaryocytes of mice transgenic for human G-CSF had a modal DNA class of 8N, showing a striking contrast to the previous reports that normal mouse megakaryocytes from most strains have 16N DNA content as a modal class. Daily 10 micrograms administration of G-CSF to mice for three to five days affected the DNA distribution pattern of bone marrow megakaryocytes, with a higher proportion of cells having 8N DNA contents. This G-CSF treatment, however, did not influence the peripheral blood platelet count or bone marrow megakaryocyte number. Administration of G-CSF along with thrombopoietin (TPO) reduced the proportion of megakaryocytes, with 32N DNA, the DNA class that was increased by TPO. Finally, the presence of mRNA for the mouse G-CSF receptor was demonstrated in two megakaryoblastic cell lines by reverse transcriptase polymerase chain reaction. These results indicated that G-CSF may have a suppressive effect on the maturation of mouse bone marrow megakaryocytes when monitored by the DNA polyploidy. Although further study is clearly necessary, the presence of mRNA for the G-CSF receptor in megakaryocytic lineage strongly suggests the direct action of G-CSF on this cell lineage.

Animals↗
PubMed2000

Apoptosis is a mode of cell death in the polykaryon-forming unit assay.

In the polykaryon-forming unit (PFU) assay, which defines cell survival as the ability to form a cytochalasin-induced polykaryon of predetermined ploidy, the mode of PFU deletion is not known. Incubation of L5178Y-S PFU in cytochalasin resulted in polyploidy (> or =32C) and most polykaryons (>75%) ultimately underwent apoptosis, detected using chromatin condensation and externalised phosphatidylserine. However, large polykaryons carrying terminal deoxynucleotidyl transferase-mediated dUTP nick end-labelling (TUNEL)-labelled DNA strand breaks were not observed, presumably due to rapid loss of DNA. Gamma irradiation of PFU prior to cytochalasin exposure caused a reduction in the frequency of highly polyploid cells (>16C), consistent with either a supra-induction of apoptosis or a reduction in the ability of PFU to reach high ploidies. We conclude that L5178Y-S PFU are deleted by apoptosis.

Animals↗
PubMed1997

Phylogeny and phylogeography of the circumpolar genus Fraxinus (Oleaceae) based on internal transcribed spacer sequences of nuclear ribosomal DNA.

The DNA sequences of the nuclear internal transcribed spacers (1 and 2) of nuclear ribosomal DNA were determined for 27 taxa of Fraxinus in an attempt to reconstruct the phylogeny of the genus and interpret its biogeographic history in a phylogenetic context. Minimal intra-individual and intraspecific polymorphisms were observed and infrageneric taxa were all recovered in phylogenetic analyses. The tree topologies estimated from parsimony and neighbor-joining analyses of one- and two-parameter substitution rates were congruent and supported by high bootstrap estimates. Indels were also found to be phylogenetically informative. The phylogenetic trees obtained were in quite good agreement with the current taxonomy of the genus but homoplasy was observed in the evolution of floral characters. Polyploidy also appeared to have evolved several times in the genus. Fraxinus nigra was more closely related to European and Asian taxa than to the other American ashes. The Asian F. platypoda had a closer relationship to Asian and European ashes in section Bumelioides than to its presumed North American allies of section Melioides. With respect to the position of F. quadrangulata as sister group to the remainder to the remaining species, the subgenus Fraxinus appeared paraphyletic. Phylogeographical analysis indicated that Fraxinus likely originated in North America, with two subsequent events of intercontinental migration from North America to Asia.

Asia↗
PubMed1993

Analysis of cell ploidy in histological sections of mouse tissues by DNA-DNA in situ hybridization with digoxigenin-labelled probes.

DNA-DNA in situ hybridization, with two digoxigenin-labelled, chromosome-specific DNA probes, was used to determine the number of copies of a given chromosome in interphase nuclei and so identify putatively polyploid nuclei in histological sections of several mouse tissues. One hybridization site per diploid genome was expected for tissues with hemizygous markers: male mice hybridized with a Y chromosome probe (pY353/B) or hemizygous transgenic mice hybridized with a beta-globin probe (pM beta delta 2). Nuclei with more than one hybridization site were considered putative polyploids. Three groups of experiments were undertaken: (1) evaluation of the method, using mouse liver sections; (2) studies of tissues already known to contain polyploid nuclei, and (3) studies that resulted in the discovery that the mouse ovary contains polyploid nuclei. First, control studies showed that the ability to detect the target DNA sequences was affected by section thickness. Studies of nuclear ploidy in the developing mouse liver revealed a pattern similar to that established by previous studies using DNA content as a criterion for ploidy. At birth, only about 5% of the liver nuclei were polyploid; this increased to 10-15% by 10-20 days and was followed by a sharp increase in the frequency of tetraploid nuclei between 20 and 40 days (to about 35%) and a more gradual increase in higher order polyploid nuclei. Secondly, this technique was used to confirm that polyploid (mostly tetraploid) nuclei were present in the bladder epithelium, heart, uterine decidua and placental trophoblast. Higher order polyploidy was seen in large bone marrow cells (megakaryocytes) but not in the even larger trophoblast giant cells of the placenta, thus confirming previous claims that these cells are polytene rather than polyploid. Thirdly, putatively tetraploid nuclei were found in the ovarian follicle and corpus luteum. As far as we are aware, this is the first time polyploid nuclei have been reported for the mouse ovary.

Animals↗
PubMed1977

An explicit formula for frequency changes in genetic algebras.

A general situation in population genetics is considered with any hereditary system described by a genetic algebra. It is assumed that there is random mating, no selection, and infinite population size. A formula is derived for the distribution of genetic types in the general jth generation given the distribution in the initial generation. Special attention is paid to the case of one locus with polyploidy and chromosome segregation. The calculations are carried out as far as possible in the situation with triploid gametes.

Gene Frequency↗
PubMed1982

The expression of antibody diversity in natural and laboratory-made polyploid individuals of the clawed toad Xenopus.

Antibody diversity, as measured by isoelectric focusing of dinitrophenol-specific antibodies, was compared in different polyploid species of the clawed toad Xenopus. Antibody heterogeneity increased with chromosome number and DNA content from Xenopus tropicalis (2n = 20 chromosome) to Xenopus ruwenzoriensis (2n = 108 chromosomes). Laboratory allopolyploids made by hybridization between two species showing different antibody diversities and different chromosome numbers gave antibody patterns intermediate between the two parents. On the other hand, autopolyploid individuals showed no increase in antibody diversity, showing that increased polyploidy alone cannot be responsible for increased heterogeneity. In contrast to the increase in antibody diversity following polyploidization, the number of expressed major histocompatibility complex alleles, as measured by a mixed lymphocyte reaction, did not increase. This locus appeared to be diploid or in the process of rediploidization in all the Xenopus species studied. Selection has thus operated differentially on the polyploid immunoglobulin and major histocompatibility loci. It apparently preserved the additional heterogeneity acquired for immunoglobulins favoring the expression of an expanded antibody repertoire in polyploid species.

Animals↗
PubMed1984

The effect of vinca alkaloids in enhancing the sensitivity of a methotrexate-resistant (L1210/R7A) line, studied by flow cytometric and chromosome number analysis.

Two L1210 murine lymphoma cell lines sensitive and resistant to methotrexate (L1210 and L1210/ R7A , respectively) and previously shown to exhibit collateral sensitivity to the vinca alkaloids have been studied by flow cytofluorimetric techniques following propidium iodide staining of the DNA. Following treatment with a range of concentrations of vincristine, both cell lines showed a build-up of fluorescence in the 4n position. However, the methotrexate-resistant cell line exhibited this effect at lower doses of vincristine. On an equimolar basis, the vinca alkaloids ranked for intensity of this effect in the order vinblastine greater than vindesine greater than vincristine. DNA fluorescent histograms following various times of continuous exposure to vincristine showed an accumulation of material at the 8n position, which was shown by chromosome analysis to be due to polyploidy. It was concluded that methotrexate-resistant cells (L1210/ R7A ) experience difficulty in traversing mitosis and this difficulty is enhanced by the vinca alkaloids.

Animals↗
PubMed1976

The ultrastructure of polyploid B-cells in the islets of normal mice.

Light and electron microscopic studies of diploid, tetraploid and octaploid B-cells in the islets of normal C57BL/KsJ mice revealed that polyploid cells were characterized by a wider range of granulated states than diploid B-cells. The maximum granule densities were similar for polyploid and diploid cells; however, some polyploid cells were almost devoid of granules, while the least granulated diploid cells contained intermediate granule densities. The tetraploid cell also appeared to be characterized by an increased mitochondrial stage which suggests compensation for the greater degree of degranulation. These observations were confirmed by morphometric analysis. Two interpretations of the apparent polyploidy are discussed; that polyploid B-cells may be more responsive to insulin releasing stimuli than diploid B-cells and that tetraploid cells may only be diploid cells in the G2 phase of the mitotic cycle.

Animals↗
PubMed1979

Alcohol dehydrogenase activity in diploid and autotetraploid Phlox.

The effect of polyploidy on the activity of alcohol dehydrogenase was examined in a series of diploid and synthetic autotetraploid Phlox drummondii. In most cases autotetraploids had about twice as much activity as the corresponding diploids, in two cases autotetraploids had about 1.5 times more activity, and in one wild seed pair the activity of the tetraploid was somewhat lower.

Alcohol Oxidoreductases↗
PubMed1986

Polyploidization and localisation of poly(A)+ RNA in the different cell types of the vitellogenic meroistic ovary of the fleshfly, Sarcophaga bullata.

The degree of polyploidization, the level of transcriptional activity and the volume of the different cell types present in the meroistic ovary of Sarcophaga bullata were measured during different vitellogenic stages. The nurse cells and the germinal vesicle exhibited very pronounced differences with regard to DNA content and mRNA synthesis, even though they are genetically identical. During the 4C stage (late vitellogenesis), we observed different degrees of polyploidy in follicle cells adjacent to the oocyte and those surrounding the nurse cells. Although the chromatin of the germinal vesicle is condensed into a karyosome, in situ hybridisation revealed the presence of transcriptional activity. The volume of the germinal vesicle, which contains only 4C DNA, is big enough to contain 2048C DNA. The meroistic ovary is a highly polarized differentiating system. Our results are discussed in the light of the fact that the polytrophic ovary is a miniature electrophoresis chamber.

Animals↗
PubMed1976

Originalarbeiten.

In 5 cases of polycythaemia vera and 2 cases with other myeloproliferative disorders accompanied by thrombocythaemia (megakaryocytic myelosis), the megakaryocytes were differentiated and studied by use of the combined application of cytophotometric determination of the DNA content and autoradiography with tritiated thymidine (3H-TdR) in vitro. A shift to the right of the megakaryocyte series, occurence of high polyploidy cells at 64c and a decrease of the 3H-TdR-labeling indices were observed. The data suggest a disturbance of the rhythmical polyploidization of the megakaryocytes, consisting of an elevated proportion of rest cells at the different ploidy stages. The maturation capacity of megakaryocytes may be related more to the resting than to the DNA synthesizing cells.

Adult↗
PubMed1976

A CHO-cell mutant with a defect in cytokinesis.

In a selection procedure designed to enrich for temperature-sensitive mutant cells blocked in mitosis a CHO-cell mutant was isolated which has a defect in cytokinesis as the basis of its temperature-sensitive phenotype. Cultures of the mutant had an abnormally high percentage (ie. 34%) of polyploid cells at the permissive temperature of 34 degress C and showed further increased frequencies of polyploidy as well as many multinucleated cells at 38.5 degrees 39.5 degrees. When the mutant cells were synchronized in metaphase by Colcemid arrest and then placed into fresh medium at nonpermissive temperature, they did not divide although the completion of mitosis appeared cytologically normal. Ultrastructural examination by electron microscopy of such synchronized cells at telophase revealed no specific defects in cellular components other than failure of development of a normal midbody. The sensitivity of the mutant to cytochalasin B and to Colcemid was the same as for wild-type cells. This mutation behaved as recessive in tetraploid cell hybrids constructed by fusing the mutant with a CHO strain which was wild-type with respect to temperature sensitivty.

Animals↗
PubMed1978

Selfing in genetic algebras.

The effect of self fertilization on the distribution of genetic types in a population can be represented algebraically by a linear transformation. In this paper the relationship of the transformation to the genetic algebra governing the population is investigated. In particular, the problems of multiple alleles, polyploidy and linked loci are studied.

Alleles↗
PubMed1976

Karyology of primary human fetal cell cultures.

Cell cultures were established from the biopsies of lung, skin and kidney from each of nine human fetuses, and chromosome analyses were performed on material through the fifth subculture. Kidney cell cultures generally showed a higher level of polyploidy than lung or skin. The frequencies of hyperdiploid cells and those with structural abnormalities were consistent with the low levels found in cultures of human lymphocytes. The data provide a normal cytogenetic baseline for human fetal material which may be useful in a variety of studies.

Cells, Cultured↗
PubMed1985

Some cell kinetic effects of combined injury with ionizing radiation and cyclophosphamide on mouse bladder urothelium.

Cyclophosphamide was given intraperitoneally to groups of eight female mice 48 h after local electron irradiation to the bladder with 0, 10 and 20 Gy respectively. The reactions in the urothelium were monitored by histology, incorporation of tritiated thymidine and flow cytometry. A wave of increased thymidine incorporation combined with an increase in the proportion of diploid S-phase cells was seen in the unirradiated bladders 24 h after the drug treatment, followed by normalization after 1 week. This response was significantly less pronounced in the irradiated animals. In the unirradiated animals a similar wave characterized by an increased proportion of octaploid cells was also seen, but this wave occurred later in the irradiated animals. Severe injury was observed in the rectum of the 20 Gy-irradiated animals. Irradiation prior to drug treatment led to only small effects, but a decreased ability for regenerative DNA synthesis after drug injury seems to persist. This affects both proliferation and the building up of polyploidy.

Animals↗
PubMed2006

Cyclin A1 is a p53-induced gene that mediates apoptosis, G2/M arrest, and mitotic catastrophe in renal, ovarian, and lung carcinoma cells.

We were the first to identify cyclin A1 as a p53-induced gene by cDNA expression profiling of p53-sensitive and -resistant tumor cells [Maxwell S. A. and Davis G. E. (2000) Proc. Natl. Acad. Sci. USA 97, 13009-13014]. We show here that cyclin A1 can induce G2 cell cycle arrest, polyploidy, apoptosis, and mitotic catastrophe in H1299 non-small cell lung, TOV-21G ovarian, or 786-0 renal carcinoma cells. More cdk1 protein and kinase activities were observed in cyclin A1-induced cells than in GFP control-induced cells. Thus, cyclin A1 might mediate apoptosis and mitotic catastrophe through an unscheduled or inappropriate activation of cdk1. Two primary renal cell carcinomas expressing mutated p53 exhibited reduced or absent expression of cyclin A1 relative to the corresponding normal tissue. Moreover, renal carcinoma-derived mutant p53s were deficient in inducing cyclin A1 expression in p53-null cells. Cyclin A1 but not cyclin A2 was upregulated in etoposide-treated tumor cells undergoing p53-dependent apoptosis and mitotic catastrophe. Forced upregulation of cyclin A2 did not induce apoptosis. The data implicate cyclin A1 as a downstream player in p53-dependent apoptosis and G2 arrest.

Apoptosis↗