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Colchicine solution was applied to the primary adventitious buds of Angelica dahurica var. formosana in vitro to induce the polyploid. Compared with non-treated plantlet, the morphology, microhisology, and chromosome number of treated plantlets are varied. It proved that the polyploid induction was effective.
A decrease in the effectiveness of neutron-irradiation with respect to fusion of nonproliferating hepatocytes of animals with age was shown by the method of flow cytometry. There was an inverse relationship between the effectiveness of induction of nonproliferating hepatocytes fusion and neutron energy. The process of hepatocyte fusion induced by neutrons was inhibited by uranyl acetate. No age-dependent changes were noted in the induction of polyploidization of proliferating hepatocytes by sparsely ionizing radiation. A hypothesis is proposed concerning a membrane nature of the target responsible for hepatocyte polyploidization induced by densely ionizing radiation.
Total ecdysones (ecdysterone and inokosterone) were isolated and identified from Achyranthes bidentata radix of polyploid and monoploid by column chromatography. The hormone contents of monoploid and diploid are about the same, but the content of polyploid is 14 times that of the diploid.
The method of flow cytometry was used to study polyploidization of hepatocytes following X-, gamma-, and neutron-irradiation. Ionizing radiation was shown to induce cell polyploidization by two different ways: (1) cells and nuclei fusion, and (2) restriction of mitosis after DNA replication. RBE of 14 MeV neutrons with respect to fusion was about 5.10(3). With neutron irradiation, the sensitivity of cells by fusion was not lower than that by chromosome mutations.
The pieces of ventricles of newborn rats, which consist of diploid myocytes in more than 95%, were transplanted under the renal capsule of a syngeneic adult rat. Within 35 days the transplants contained 40 to 60% of the cells with the double or even greater DNA content in the mononuclear and binuclear classes. The composition of the classes was similar unlike the number of the cells of a certain class in the transplant and in the heart of a 35 day old rat. A conclusion has been drawn that the polyploidization is programmed and the realization of this programme depends on the growth, work, and other factors but is expressed even at their minimal action in the transplant. The growth of the transplanted myocytes was weaker than in the heart. It is supposed that, unlike the polyploidization, the growth of the myocytes outside the cycle is, predominantly, functional-dependent.
Methodological approaches to kinetics of cell polyploidization in the rat liver parenchyma are discussed. Different ways of hepatocyte polyploidization in the course of postnatal liver growth have been assessed. The intensities of hepatocyte transitions from one ploidy class to another were determined. On the basis of literary experimental data the following is summarized: With the increase in the animal age, there is a decrease in hepatocyte transition from one ploidy class to and ther; in young animals the intensity of formation of tetraploid hepatocytes through the stage of binuclear cells (2c----2c X 2----4c) is 0.39-0.55 within two weeks, the intensity of direct transitions (2c----4c) being 0.00-0.19 within the same time. The intensity of entering to DNA synthesis is reduced with the increase in hepatocyte ploidy levels; in this case the coefficient of the reducing of mitotic activity is calculated as 0.10-0.22, and 0.01-0.05 for 4c- and 8c-hepatocytes, resp. The factors stimulating proliferation in the liver increase the intensity of the direct cell transition (2c----4c) by several times which can exceed the intensity of transition through the binuclear cell stage.
Two pyrimidine analogs (metahalones) NY 3163, NY 3170 have been tested for their effects on the growth of rat glioma cells in monolayer and in spheroid culture. Both substances have earlier been found to inhibit the microtubule system in malignant cells. In glioma cells, arrest of mitosis was accompanied by repeated cycles of DNA synthesis, leading to different levels of polyploidization up to 16 and 32 ploid cells. The effect was not reversible through a culture period of 4 days. Reduced ability of directional migration of cells on a plastic surface was seen for 16 days after exposure. The reaction to microtubule inhibitors seems to differ depending on the type of tumour cells, where some malignant cells have earlier been reported to escape mitotic arrest and proceed to the next G1 phase, in contrast to the present glioma cells which undergo polyploidization.
In order to see whether the polyploid cells lining the mouse urinary bladder are formed by nuclear fusion, such epithelium was studied under the light and electron microscope forty-eight hours after an injection of cyclophosphamide when the bladder epithelium regenerates with rapid formation of many diploid, tetraploid and octoploid cells. The probability of seing fusion, if it occurs, ought then to be high. Serial sections of many specimens from four mice revealed no signs of fusion. Thus we found no support for the theory that polyploid cells are formed by nuclear fusion.
The intranuclear mitosis of the plasmodial nuclei of myxomycetes permits the observation of defects in chromosomal repartition which would probably be lethal in other eukaryotic cells with open mitosis. We found that antitumoral platinum-amine compounds perturbed late mitotic events and induced the formation of giant nuclei which were polyploid in plasmodia of Physarum polycephalum. Using 26 platinum-amine complexes, we have shown that all antitumoral compounds induced the formation of polyploid nuclei for drug concentrations at least three times lower than the amount necessary to block the overall plasmodial growth, whereas platinum compounds without antitumor activity did not behave this way. DNA replication appeared to be quantitatively normal during formation of giant nuclei by antitumoral compounds. These observations suggest that platinum-amine compounds exert their antitumor activity by interfering with mitosis rather than by a gross inhibition of DNA synthesis.
Three nearhexaploid sublines were obtained from hypotriploid mouse L cells by means of colcemid treatment. When cultivated on solid substratum, all of them did not differ from the parental line either in doubling time or in cloning efficiency. The ability of polyploid cell variants to be initiated for proliferation in a semi-solid medium was equal to that of hypotriploid cells, while the average diameter of colonies formed by hexaploid cells in methyl cellulose turned out to be significantly smaller than the size of colonies of parental cells. The inhibition of growth in the semi-solid medium may reflect partial normalization of the transformed phenotype of polyploid L cells.
Ventricular cardiomyocytes of 5-6 day-old mice are mostly polyploid. At this time 60% of the cardiomyocytes become binuclear and another 10% mononuclear polyploids. The binuclear cardiomyocytes emerge as a result of acytokinetic mitosis, while mononuclear tetraploids as a result of the cessation of the mitosis at the initial stages.
Polyploidization of hepatocytes in albino rats subjected to continuous and intermittent exposure to CCl4 has been studied karyometrically. It has been shown that polyploidization as a response to the exposure of chlorderivatives of hydrocarbons is determined by hepatotoxin toxicity related to the number of chlorine atoms in the molecule and to the character of the exposure involving route, dose and regimen of exposure.
Near DNA diploid human adult solid tumors are often associated with certain near-tetraploid cells. In an established human breast cancer cell line, Hs578T, with a DNA index in the hyperdiploid region, polyploid cells appeared during exponential growth. Among clones generated from single cells and analyzed by the video time lapse technique, an intraclonal interdivision time (IDT) heterogeneity is presented that renders endoduplication a plausible explanation for the generation of the polyploid cells observed. This conclusion, drawn from our IDT analysis, is supported by curves drawn from counting grain-positive cells during continuous labeling with [3H]-thymidine. Our results are compared with a parallel analysis of the aneuploid human breast cancer cell line MDA-231, generating intraclonal IDT heterogeneity, due mainly to the mitotic instability of that line, as we reported previously.
Mitotic figures in the mink placental trophoblasts have been observed under the light microscope using actions and air-dried preparations. The tetra- and octaploid metaphase chromosome spreads were found on Giemsa-stained air-dried preparations. A high percentage (up to 80%) of abnormal metaphases, including k-mitoses as well as a portion of restitution anaphases, was revealed on sections of the placental trophoblast suggesting a possible block of mitosis at the meta- and anaphase. Therefore, it is very likely that genome multiplication in a portion of placental trophoblast cells in mink involves block of mitoses at meta- and anaphase followed by restitution. The chromosomal arrangement on metaphase spreads in part of cells showed a degree of separation of the whole di- and tetraploid chromosome sets within tetraploid and octaploid chromosome plates. Several spreads exhibited some allocycly of diploid chromosome sets inside the polyploid metaphases. It is not inconceivable that such an arrangement may reflect some autonomy of low-ploidy chromosome sets within the polyploid trophoblast cells in mink.
In this study, we analyzed effects of copper toxicity on the process of mitosis by examining polyploid accumulation and histone H1 hyper-phosphorylation, which result from defects in mitotic completion, in regenerated LEC rat liver. The proportion of polyploids with 8c nuclear DNA content in LEC rats was significantly higher than that in normal rats (P < 0.05). The degree of histone H1 phosphorylation was about 2.5-fold higher in LEC rat than normal rat. These results suggest that the accumulated copper in LEC mutant rat causes the block of mitotic completion in cell cycle.
Levels of hepatocyte ploidy were investigated in patients with chronic viral hepatitis, alcohol hepatitis, and liver cirrhosis. It was shown that in spite of some decrease in the share of mononucleate diploid hepatocytes these cells equally make the main part of hepatocyte population (about 80% on the average) both during chronic liver diseases and in the norm. In chronic hepatitis and cirrhosis processes of polyploidization in the human liver, unlike those in livers of laboratory animals (rats and mice), are accomplished with much lower rates and characterized by accumulation of binucleate diploid cells. These processes differ from those occurring under ageing. Reparative regeneration of human liver under chronic hepatitis and cirrhosis is accomplished on the whole by proliferation of mononucleate diploid hepatocytes but processes of polyploidization in the liver of some patients can play a significant role in reparative liver growth.
This ultrastructural study of endomitosis in polyploid albumen gland cells of succineid snail. S. lauta was made to extend the previous research (Anisimov, 1997). The investigation was prosecuted on ultrathin sections by means of transmission electron microscopy. The insertion of 3H-thymidine and 3H-uridine into the nuclei and the light microscopy control of nuclear ploidy allowed to identify glands on early and intensive polyploidization stages and to ignore pseudoendomitotic terminally differentiated cells. Considering ultrastructural qualities and information about the cell population kinetics enabled us to discriminate between predifferentiation (2-4c), protodifferentiation (4-8c) and promoted differentiation (8-16c) cell stages. The endomitotic cycle was observed synchronously with the development of cytoplasmatic structures and secretion production in differentiating cells of ploidy levels 4-8-16c. According to morphological condition of the chromatin and chromosomes, G1-, S- and G2-interphase periods can be determined, such as successive stages of endomitosis (endopro-, meta-, ana-, and telophase). The following main features of endomitosis are accentuated: the normal (mitotic) chromosome cycle with total compaction in endometaphase and differential decompaction in interphase; the absence of the mitotic spindle; integrity of the nuclear envelope, nucleolus and contacts between them and chromosomes; part of chromosomes split nonsimultaneously and incompletely in the endoanaphase, or endomitosis with diplochromosomes rarely occurs. Chromosome nondisjunction is a temporary process that does not lead to a permanent chromosome endoreduplication (polyteny). It is emphasized that the normal chromosome cycle is combined with a complete block of cytoplasmic structures. Possible reasons of mitotic cycle reduction during the transition of differentiating cells to endomitotic polyploidy are discussed.
Using a mature albumen gland of Succinea lauta, a study was made of the chromatin structure and RNA synthesis activity in polyploid nuclei (16c-32c-64c) of terminally differentiated cells, by means of electron microscopy and 3H-uridine radioautography of squash preparations. No 3H-thymidine incorporation in these cells was observed, which suggested the absence of DNA synthesis and endomitotic cycles at the examined functional stage of gland development. Some different stages of secretory cells could be distinguished: a resting period of the terminally differentiated cells, with the least 3H-uridine incorporation; a phase of intense heterosynthetic transcription, with the highest 3H-uridine incorporation; intervals in daily cycles; nuclear degeneration in ageing cells. At all these states the nuclei demonstrated some morphological features of endomitosis. This may be possible, because in the course of terminal cell differentiation the main chromatin mass is compacting like chromatids or chromocentres with thin euchromatinous filaments, which can be covered with perichromatinous granules. Since the true endomitosis is deliberately excluded at this stage, the above described morphological state of polyploid nuclei is regarded as postendomitotic and is designated as pseudoendomitosis. The view of the existence of "two fundamentally different types of endomitosis", one of which is "endomitosis in general", and the other "shows up as stationary" (Therman et al., 1983), is accepted as groundless.