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PubMed1979

Functional anatomy of haploid and diploid rabbit spermatozoa.

Ultrastructural studies of surface replicas of haploid and diploid rabbit spermatozoa after treatment with various chemical media have shown that the various structural components of the diploid sperm head possess the same stabilities and labilities as those of the haploid. Therefore, at least in terms of functional anatomy, diploid rabbit spermatozoa should be capable of penetrating the egg investments and undergoing syngamy.

Acrosome↗

PubMed2003

Phylogenetic relationship of ribosomal ITS2 and mitochondrial COI among diploid and triploid Paragonimus westermani isolates.

We compared patterns of intraspecific polymorphism of two markers with contrasting modes of evolution, nuclear ribosomal DNA (rDNA) and mitochondrial DNA (mtDNA), in the lung fluke, diploid and triploid Paragonimus westermani from three geographical regions of Korea. The genetic distances between three populations of Korean diploid and triploid P. westermani showed no significant difference in the nucleotide sequences of the mitochondrial cytochrome c oxidase subunit I (mtCOI) and ribosomaal second internal transcribed spacer (ITS2) genes. A highly resolved strict-consensus tree was obtained that illustrated phylogenetically useful information of the ITS2 and mtCOI sequences from diploid and triploid P. westermani.

Animals↗

PubMed1999

Maintenance of near-diploid karyotype of PA-1 human ovarian teratocarcinoma cells due to death of polyploid cells by chromosome fragmentation/pulverization.

Chromosome instability (polyploidy or aneuploidy) is one of the characteristics of malignant tumors. Human teratocarcinoma cell line PA-1, which was established more than 10 years ago, consists of a majority of near-diploid cells and a minority of polyploid cells, indicating that it is karyologically very stable. In the present study we investigated this genomic stability from the view point of cytogenetics. Cleavages and breaks in the chromosome were found in the metaphase of PA-1 polyploid cells, accompanied by the formation of polynucleosomal DNA fragments. These findings were absent in the near-diploid cells. In addition, polyploid cells did not show colony-formation ability by in situ analysis of cytogenetics in each colony. Thus, the maintenance of the near-diploid karyotype in PA-1 cells may be due to a blockage in the M-phase of the polyploid cells by functional mitotic checkpoints, if any, leading to cell death due to inability to enter the next cell cycle.

Chromosome Banding↗

PubMedDate not supplied

Reappraisal of karyological monitoring of diploid cell substrates.

The practical aspects of present requirements for karyological monitoring of diploid human fibroblasts used in vaccine production are critically re-examined. A brief review of known chromosomal instabilities and their correlations with neoplasia and with in vitro conversions of cell growth is presented and discussed in relation to karyological criteria in the selection of mammalian diploid cell substrates. It is proposed that extensive experience with the growth and karyological characteristics of human diploid fibroblast cell strains of embryonic origin may warrant the reduction or elimination of karyological monitoring of production lots. Additional requirements for characterization of cell strains for approval as substrates for vaccines are suggested which involve evaluations of chromosomal stability based upon (a) viral transformation rate and (b) aberration production rate in response to x-irradiation.

Cells, Cultured↗

PubMedDate not supplied

The development of rabbit diploid cell lines.

The authors describe briefly studies on the development of diploid cell lines from fetal tissues (origin: four inbred rabbits). Some lines have been maintained in continuous serial culture for over three years. The Lung 16 may represent a qualifying rabbit diploid cell line when used between cell passages 48 and 100; during this period it has shown a minimum deviation from the normal diploid rabbit karyotype; it supports the growth of rubella virus, is non-tumorigenic and might be useful as a substrate for human vaccine.

Animals↗

PubMed1975

Growth and death of diploid and transformed human fibroblasts.

Three possible explanations are presented for the differences in growth potential between human diploid fibroblasts of finite life-span and permanent transformed lines: 1) Only diploid cells have a molecular clock mechanism which counts cell divisions prior to senescence. Two hypothetical examples of such mechanisms are described; however, the available evidence argues against a clock mechanism for aging in fibroblasts. 2) Cells become committed with a given probability to a slow buildup in protein errors, which leads after many divisions to a lethal error catastrophe. It can be shown that speeding up the rate at which the error catastrophe develops, as may occur in transformed cells, can convert a population of finite life-span to one with infinite growth. 3) The growth rate of diploid cells may not depend on the limiting concentration of any one protein. If so, cells with a low level of errors will not have a reduced generation time, and there will be no selection against them. On the other hand the uncontrolled growth of transformed cells may be reduced in rate by the presence of faulty proteins, so that there is continuous selection for those with the fewest errors. Finally, the analogous problem of the mortality of somatic cells and the immortality of the germ line is also briefly discussed.

Animals↗

PubMed2000

Re-analysis by fluorescence in situ hybridisation of spare embryos cultured until Day 5 after preimplantation genetic diagnosis for a 47, XYY infertile patient demonstrates a high incidence of diploid mosaic embryos: a case report.

Mosaicism in 4-8-cell human embryos analysed by fluorescence in situ hybridisation (FISH) has been widely reported, but few studies have addressed the incidence of mosaicism in more advanced embryonic stages. In the present study we analysed spare human embryos in a case of preimplantation genetic diagnosis (PGD) for increased risk of aneuploidy because of an infertile 47,XYY man. After replacement of two embryos typed as 1818XX at PGD, six spare embryos (not frozen because of their low quality) were re-analysed on Day 5 for PGD confirmation. Out of five embryos typed as 1818XY at PGD, four were diploid mosaic (DM) and one was normal in all cells. The sixth embryo, typed as 18XYY/1818181818X at PGD, was a DM. In spite of the bias of our small series of morphologically low-quality embryos, the surprisingly high proportion of mosaics (which confirms previous findings) questions the validity of PGD, but supports the strategy of transferring only the embryos where two blastomeres gave normal and concordant results at PGD. More data are required to understand the clinical significance of early diploid mosaicism (and its impact on implantation rate) and to determine whether some diploid mosaic embryos might be considered safe for transfer.

Adult↗

PubMed2001

Mouse singletons and twins developed from isolated diploid blastomeres supported with tetraploid blastomeres.

The aim of this study was to obtain mice, hopefully identical multiplets, from single diploid blastomeres isolated at the 4-cell stage, or from pairs of sister blastomeres isolated at the 8-cell stage. To this end isolated blastomeres were aggregated with one or two tetraploid carrier embryos produced by electrofusion of 2-cell embryos. Diploid embryos were albino and homozygous for the "a" allele of glucose-phosphate isomerase (GPI-1a1a) and tetraploid embryos were pigmented and GPI-1b1b. The aggregates were cultured in vitro up to the blastocyst stage. Each quartet (occasionally triplet or doublet) of chimaeric blastocysts was transplanted to the oviduct of a separate pseudopregnant recipient. Altogether 62 blastocysts were transplanted to 17 recipients. Eight full-term foetuses (two singletons and three pairs of twins) were rescued by Caesarian section on day 19, 20 or 21 of pregnancy. Three young (one singleton and twins) were successfully reared by foster mothers and proved to be normal and fertile females. All foetuses and animals were albino. In five individuals only the 1-A form of GPI (characteristic for 2n blastomere) was found. In one adult female traces of the 1-B form of GPI (characteristic for 4n carrier blastomeres) were detected in the heart and the lungs while 4 other organs contained only the 1-A form. These observations strongly suggest that the majority of foetuses/animals produced according to our experimental system are 'pure' diploids rather than 2n/4n chimaeras, and that the described method can be used in future to produce twins, triplets and quadruplets in the mouse. Our study confirms earlier work by Kelly (1975, 1977) that 'quarter' blastomeres of the mouse are still totipotent.

Animals↗

PubMed1975

[Ribosomal RNA synthesis in haploid and diploid loach embryos].

rRNA synthesis was compared in the loach haploid (In) and diploid (2n) embryos. The relative intensity of synthesis was evaluated by 14C-uridine incorporation in 27S and 18S rRNA isolated from ribosomes taking into account label incorporation into total acid-soluble fraction and phosphrylated uridine derivatives. Label incorporation into rRNA, in reference with DNA content in 1n and 2n embryos, suggests that the level of rRNA synthesis per DNA unit in haploids is twice that in diploids whereas, in reference per cell, the same amount of ribosomes is synthesized both in haploids and diploids. The data obtained show that the amount of rRNAs synthesized in the loach embryogenesis does not depend on ploidy.

Animals↗

PubMed2002

[Meiosis spindle formation and chromosome behavior in diploid potatoes with 'fused spindles' mutation].

Chromosomal behaviour and spindle morphology were studied in microsporogenesis of two kinds of diploid potato clones: with normal meiosis, and with "fused spindles" (fs) occurring during the second meiotic division from prometaphase II (proMII) to telophase II (TII). For the first time, morphological effect of fs was found at the late proMII stage to be expressed as two interrelated processes: 1) abnormal chromosome movement, which resulted in joining two groups of chromosomes in the central zone of meiocytes, and 2) abnormal formation of two spindles in the direction to two division poles instead of four poles that actually led to the formation of a united bipolar spindle. Thus, it is not the fusion of two parallel spindles but the formation of united bipolar spindle that constitutes fs abnormality, while the parallel co-orientation of two spatially separated meiotic spindles is a norm in diploid potato. These primary abnormalities detected at proMII resulted in abnormalities at its subsequent meiotic stages: formation of fused spindle and united metaphase plate at MII, bipolar chromosome segration at anaphase II, formation of two telophase nuclei at TII and dyads at the tetrad stage. The results obtained evidence the polar division disturbance in diploid potato clones with fs abnormality.

Chromosomes, Plant↗

PubMed1977

The repair of X-ray induced chromosomal damage in trisomy 2-and normal diploid lymphocytes.

The frequency of chromosomal aberrations produced by X-rays is greater in lymphocytes cultured from trisomy 21 patients (Down's syndrome) than from normal diploid donors. This increase, which can be detected by a micronucleus assay for chromosomal damage, was postulated by us to result from a defect in the rejoining system which repairs chromosomal breaks. The postulated defect would result in a longer rejoining time, therapy permitting more movement of broken ends and thus enhancing the frequency of exchanges. To test this possibility, the time required for the rejoining (repair) of chromosome breaks was measured in lymphocytes from five Down's syndrome (four trisomy 21 and one D/G translocation partial trisomy 21) donors, from a monosomy 21 donor, and from five diploid donors. The rejoining time was reduced in the Down's syndrome lymphocytes in comparison to the normal diploid and monosomy 21 lymphocytes. Thus the repair of chromosome breaks, far from being defective as evidenced by a longer rejoining time in Down's syndrome cells, occurred more rapidly than in normal cells. A mechanism is proposed by which reduced rejoining times would increase aberration frequencies as a consequence of competition between a (hypothetical) error-free repair system and the error-prone repair system that generates chromosomal aberrations. We suggest that the alteration in the rejoining of chromosomal aberrations may underlie the increased susceptibility of people with Down's syndrome to leukemia.

Cells, Cultured↗

PubMedDate not supplied

[The development of diploid parthenogenetic mouse embryos of the inbred C57BL and CBA strains].

We studied preimplantation development in vitro and postimplantation development in vivo of diploid parthenogenetic mouse embryos of C57BL/6 and CBA strains, as well as of (CBA x C57BL/6)F1 hybrids. Development to blastocyst stage of diploid eggs obtained from C57BL/6, CBA, and hybrid mice was observed in 90, 15, and 73% cases, respectively. After implantation, C57BL/6 embryos did not develop to somite stages, while CBA and hybrid embryos reached various stages of somite formation in 45 and 30% cases, respectively. Cultivation of embryos beginning from one-cell stage in the medium containing 2% newborn calf serum increased the yield of blastocysts from 15 to 59% in CBA embryos and from 73 to 90% in hybrids; However, such effect was not observed with C57BL/6 embryos. The latest stages of development observed in CBA and hybrid diploid parthenogenetic embryos were 33-35 somites and 25-30 somites, respectively. Imprinting patterns in chromosomes of CBA and C57BL/6 gametes are discussed.

Animals↗

PubMed2003

Coexistence of triploid and diploid forms of spined loach, Cobitis taenia: a model-based approach.

A tritrophic model of Lotka-Volterra type was developed to describe the dynamics of a unisexual-bisexual complex in fish. The model assumes two genetically isolated forms, bisexual (diploid) and unisexual (triploid), competing for a common resource. The efficiency of food conversion into offspring for the unisexual form is dependent on the availability of the diploid males. By means of isoplane analysis the conditions of a non-trivial equilibrium existence are determined. These conditions imply that the unisexual form should be more fit than the bisexual one in terms of lesser mortality and/or more efficient resource utilization. During field work in the summer of 2001, a unisexual-bisexual complex of spined loach, Cobitis taenia, was found in the Sura River (Mordovia). Analysis of demographic and tropho-dynamic parameters of diploid and triploid forms in this complex suggests that the triploid form, though it does not have the presumed advantage in population fecundity, does have an advantage in survival and consumption rate. The overall ratio between population parameters of the two loach forms indicates that the theoretical condition derived from the model equations is met. Thus, ecological traits of the unisexual and the bisexual forms of spined loach make it possible for them to coexist even under conditions of fully overlapping niches.

Adaptation, Physiological↗

PubMedDate not supplied

[Competition of isogenic haploid and diploid cells of the yeast Saccharomyces cerevisiae and Pichia pinus growing in mixed populations].

During "quasi-continuous" cultivation in rich and minimal media diploid yeast cells of Saccharomyces cerevisiae completely displace isogenic haploid ones. When Pichia pinus are cultivated in the minimal medium, the diploids also have an advantage over isogenic haploids. The results are discussed within the framework of the hypothesis of fixation of diploid phase in the course of biological evolution.

Diploidy↗

PubMed1990

Age-dependent decrease in the heat-inducible DNA sequence-specific binding activity in human diploid fibroblasts.

In order to gain a better understanding of the molecular mechanism of the attenuated heat shock response in aging diploid fibroblasts (Liu, A. Y.-C., Lin, Z., Choi, H.-S., Sorhage, F., and Li, B. (1989) J. Biol. Chem. 264, 12037-12045), we examined the regulation of a heat-inducible, heat shock element (HSE) sequence-specific binding protein in IMR-90 diploid fibroblasts. Using gel retardation assay, we showed that the HSE binding activity in extracts of IMR-90 cells was very dependent on heat shock of the cells; that the induction was transient with a maximal increase observed at 1 h of heat shock. Significantly, the level of this heat-inducible HSE-binding activity was age-dependent, being high in young cells and low in old cells. By Scatchard analysis, we determined that this difference in HSE binding in young and old cells was not due to a change in the affinity of the binding, rather the level of the heat-inducible HSE-binding activity was different. The equilibrium dissociation constants (Kd) were estimated to be 30, 25, and 31 pM for cells with population-doubling levels of 22, 35, and 45, respectively; the corresponding Bmax values were 0.087, 0.042, and 0.0059 pmol of 32P-HSE bound per mg of whole cell extract protein. The decreased 32P-HSE-binding activity in the old cells was not a reflection of global alteration of many transcription factors; assay of the DNA binding activity of activating transcription factor showed little difference as a function of age. Experiments of mixing extracts from young and old cells provided evidence of a dominant inhibitor of the HSE-binding activity in old cells. Probing of protein blots with 32P-HSE showed that the subunit molecular weight of the HSE-binding protein was 83,000 in both young and old cells. The pattern of regulation of this HSE-binding protein upon heat shock and cell aging as well as the identity, in DNA sequence specificity and subunit molecular weight, of this protein with that of the human heat shock gene transcription factor suggest that the HSE-binding protein is involved in the transcriptional activation of hsp genes in IMR-90 cells. We concluded that there was an age-associated decrease in the heat shock gene transcription factor DNA-binding activity and that this could account for the attenuated heat shock gene expression in aging diploid cells.

Base Sequence↗

PubMed1990

Diploidization of bovine oocytes matured in vitro and parthenogenetically activated by electric shock.

Bovine in vitro matured oocytes were parthenogenetically activated by a single pulse of direct current. Suppression of second polar body extrusion by 60 V DC-pulse followed by cytochalasin B treatment or 240 V DC-pulse were used to diploidize parthenogenones. Analysis of the cytological events 20 h postactivation clearly documented high efficiency of both diploidization techniques. All oocytes exposed to a single 60 V DC-pulse and then to cytochalasin B for 3 to 6 h were activated and 70% to 88% possessed two pronuclei. Exposure to 240-V DC-pulse for 10 and 20 microseconds activated all oocytes, 72% and 62% formed two pronuclei, but a high incidence (22% and 30%) of degenerated oocytes was observed. Prolonged in vitro culture of oocytes after parthenogenetic activation and diploidization treatment showed that formed pronuclei were able to fuse and single prometaphase to telophase mitotic cleavage figures developed in all oocytes fixed 28 h after activation, except one cytochalasin B-treated oocyte with two prometaphase sets of chromosomes.

Animals↗

PubMed1989

Kinetics of herpes simplex virus photoinhibition by haematoporphyrin in both diploid and heteroploid cells.

The kinetics of inhibition of herpes simplex virus type 1 (HSV 1) on both diploid (CEF) and heteroploid cells (HEp2) by light-irradiated haematoporphyrin (HP) was studied. The inactivation of HSV1 by HP was drug-dose dependent and light-irradiation dependent; the viruses grown in heteroploid cells being in all cases more sensitive to inhibition than viruses grown in diploid cells. Cell toxicity by HP was markedly more evident on HEp2 cells than on CEF. The highest viral sensitivity to photodynamic inactivation by HP was found to be between the 4th and the 5th hour after cell infection, when the viral DNA synthesis is at its peak and before it is incorporated into complete virions. Microfluorometric and spectrofluorometric assays revealed that virus infected cells always take up more HP than uninfected cells, and heteroploid cells incorporated more HP than diploid cells. The possibility that an increased uptake of HP and modifications of the cell micro-environment in virus infected cells could account for the viral-inhibiting properties of HP, is discussed.

Animals↗

PubMedDate not supplied

[The ratio of collagens type I and III in cultures of human fibroblasts with diploid and aberrant chromosome complement].

The ratio of collagens I/III was studied in human diploid, trisomic and triploid fibroblast strains. The following material was used in experiments: 5 strains obtained from human spontaneous abortuses (trisomy 7, trisomy 9, triploidy), 6 diploid embryonic strains, 4 strains from patients with Down syndrome, 3 strains from healthy children, 4 strains from healthy adult donors and 3 strains from very old persons (81-107 years old). Various types of collagen were separated by means of electrophoresis. Distinct ontogenetic alterations were not observed in content of collagen III in diploid and aneuploid human cultivated fibroblasts. However, considerable alterations in the ratio of collagens I/III correlated with genetic pathologies only in the patients examined.

Cells, Cultured↗