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PubMed2001

Ribosomal RNA gene expression and chromosome aberrations in bovine oocytes and preimplantation embryos.

This review focuses on the key features of development of the bovine oocyte and embryo, with comparisons of the developmental characteristics of embryos produced in vivo and in vitro. The oocyte is transcriptionally quiescent in the primordial and primary follicle. In the secondary follicle transcription is initiated in the oocyte and a ribosome-synthesizing nucleolus is established in this cell. Transcription and nucleolar activity are enhanced in the tertiary follicle during oocyte growth. When the oocyte reaches approximately 110 microm in diameter, corresponding to a follicle of about 3 mm in diameter, transcription ceases and the nucleolus is inactivated, forming a dense spherical remnant. During the final phase of follicular dominance this remnant becomes vacuolated and, in conjunction with resumption of meiosis, it disperses. The rRNA genes are apparently re-activated during the four-cell stage, that is, the third cell cycle after fertilization, but a nucleolus is not formed. During the subsequent cell cycle, that is, during the eight-cell stage, ribosome-synthesizing nucleoli are again established. Bovine embryos produced in vitro apparently display the same pattern of nucleolus development as that in embryos developed in vivo. Examination of the ploidy of embryonic cells using fluorescence in situ hybridization has revealed that the production of bovine embryos in vitro is associated with increased chromosome aberrations in the embryos. Blastocysts produced in vitro display a significantly higher rate of mixoploidy, that is, when the embryo consists of both normal diploid and abnormal polyploid cells, than that in embryos developed in vivo. The rate of mixoploidy among embryos produced in vitro increases with increasing developmental stage. Moreover, after fertilization in vitro, initially there is a high rate of 'true' polyploidy, that is, when all cells of the embryos are polyploid. However, the polyploid embryos are eliminated before they cleave beyond the eight-cell stage, the stage at which major activation of the embryonic genome occurs in cattle.

Animals↗
PubMed1980

Resistance patterns of Walker carcinosarcoma 256 and other rodent tumors to cyclophosphamide and L-phenylalanine mustard.

Comparison is made of the development of resistance to cyclophosphamide (CPA) and L-phenylalanine mustard (L-PAM), of cross-resistance, and chromosome counts, in Walker 256 (W256), rat sarcoma R3 (R3), leukemia L1210, and Ridgway osteogenic sarcoma. For development of resistance the single maximum tolerated doses of CPA or L-PAM were used, each for two sublines in the four tumors. In W256 after only one to five treatment generations, all sublines were resistant, whereas only by generation 10 had R3/CPA, R3/L-PAM, and L1210/CPA reached marked resistance, and L1210/L-PAM reached moderate resistance. All four Ridgway osteogenic sarcoma sublines were essentially still as sensitive as the parent tumor. Long-established resistant sublines from previous studies (greater than 20 treatment generations) were used for cross-resistance, chromosome, and stability studies. All W256-resistant sublines were cross-resistant to CPA, L-PAM, and thiotepa; but the sublines of the other tumors, although showing marked, or in the case of L1210/CPA, complete resistance to their respective inducing agents, retained moderate-to-full sensitivity to the other alkylators. W256/CPA and W256/L-PAM were mainly polyploid (greater than 80% of cells), whereas the other tumors were mainly diploid or near diploid. During 10 to 20 untreated generations the degree of drug resistance remained unchanged in W256 and L1210 lines, but was reduced in R3 and Ridgway osteogenic sarcoma lines. The resistance pattern of W256 appears to be compatible with a simple selection mechanism, whereas those of the three other tumors suggest involvement of multiple determinants. This study suggests that some, but not all, tumors have universal cross-resistance between different types of alkylating agents.

Animals↗
PubMed2005

Ductal breast carcinomas with whole chromosome gains as a particular subset of near-diploid tumors with different metastasis free survival.

We recently proposed the existence of a subtype of slightly hyperdiploid ductal breast cancers with cytogenetic alterations differing from those usually observed in the majority of these tumors. We aimed to establish whether these tumors, which represent about 50% of those with a DNA index (DI) comprised between 1.1 and 1.3, correspond to a particular clinicopathological entity. A retrospective study of 1771 patients operated for ductal carcinomas was performed. Three classes of tumors constituted according to DI were compared for the usual clinicopathological factors and clinical outcome. About 690 tumors (39%) were diploid/hypodiploid (DI < 1.1), 134 (7.6%) were hyperdiploid (1.1 < or = DI < 1.3) and 947 (53.4%) were polyploid (DI > or = 1.3). Median follow-up time was 106 months (range 1-177). Polyploid tumors were significantly associated with large tumor size, advanced clinical stage, high histological grade and S-phase fraction (SPF), positive lymph nodes and loss of steroid receptors. Hyperdiploid and diploid/hypodiploid tumors were similar for all the variables except SPF which was significantly higher in hyperdiploid tumors (p < 0.001). Overall survival was similar in hyperdiploid and diploid/hypodiploid tumors in univariate and multivariate analysis, while hyperdiploid tumors were significantly related to a poorer metastasis free survival, both in univariate (p = 0.023) and multivariate analysis (p = 0.031). Despite very close initial clinicopathological and biological characteristics, hyperdiploid tumors differed from diploid/hypodiploid tumors by a higher risk of metastasis, possibly related to their increased SPF.

Aneuploidy↗
PubMed1976

[Growth of the eye and pigment epithelium of the retina during postnatal development of rats].

By the weight changes, the rat's eye grows during the whole life, whereas its scleral sector with the area equal to that of pigment epithelium of the retina grows most intensively between the 2nd and 5th days after birth. During this period, its weight attains half of the value characteristic of the scleral sector for one year old rats. This period of growth of the scleral sector coincides with the previously established peak of proliferative activity and appearance of first binuclear cells in the pigment epithelium. From the 5th day till the 12th month after birth, the weight of scleral sector increases twice and its area 6 times. This suggests that the mechanical tension of the scleral sector walls is one of the factors of growth of this eye part. On the basis of comparison of the scleral sector growth, changes in proliferative activity and number of polyploid cells in the pigment epithelium of the central zone of fundus oculi, the following periodization of the life cycle of cell population of the pigment epithelium is proposed: (1) from birth till the 15th day--period of principal determination (the number of binuclear tetraploid cells attains 80%), (2) from the 15th day till, at least, the 5th month--period of stabilization, (3) after 5 months--period of senescence characterized by the accumulation of highly ploid tri- and tetranuclear cells; its lower limit is not clearly defined.

Aging↗
PubMed2003

Chronological and replicative lifespan of polyploid Saccharomyces cerevisiae (syn. S. pastorianus).

Chronological lifespan may be defined as the result of accumulation of irreversible damage to intracellular components during extended stationary phase, compromising cellular integrity and leading to death and autolysis. In contrast, replicative lifespan relates to the number of divisions an individual cell has undertaken before entering a non-replicative state termed senescence, leading to cell death and autolysis. Both forms of lifespan have been considered to represent models of ageing in higher eukaryotes, yet the relation between chronologically and replicatively aged populations has not been investigated. In this study both forms of lifespan have been investigated in Saccharomyces cerevisiae (Syn. S. pastorianus) to establish the relationship between chronological and replicative ageing.

Barbiturates↗
PubMed1994

One of two growth hormone genes in coho salmon is sex-linked.

Salmonid fishes have two growth hormone genes resulting from their polyploid ancestry. We used the polymerase chain reaction to examine genetic variation in the third intron (C) of both of these genes in coho salmon (Oncorhynchus kisutch). A polymorphism in the length of intron C in GH-1 is due to a variable number of copies of a 31-nt repeat that is absent from GH-1 of the closely related chinook salmon (Oncorhynchus tshawytscha) and rainbow trout (Oncorhynchus mykiss). Thus, this tandem repeat sequence has become established in the genome of coho salmon since the separation of this species from its closest relatives. All male coho salmon examined have an allele at the second growth hormone gene, GH-2, that is not found in females. GH-2 is thus on the sex chromosome and there is no recombination between GH-2 and the sex-determining locus (SEX). Sequences of intron C indicate much greater divergence between the X chromosome-specific allele and the Y chromosome-specific allele within coho salmon than between the X chromosome-specific alleles of coho and the closely related chinook salmon. Thus, absence of recombination between GH-2 and SEX apparently predates separation of these two species.

Alleles↗
PubMed1991

Effect of dietary restriction and aging on polyploidy in rat liver.

Liver polyploidy levels were compared as a function of age and diet in male Fischer 344 rats between 1 and 24 months of age. Dietary restriction was imposed on one group by reducing their food intake to 60% of ad libitum food intake. Histological sections of the livers of animals at each age and diet were examined. Diploid, tetraploid and octaploid nuclei were observed, and their size and frequency established. There were no differences in the diameter or volume of these size classes as a function of age or diet. An age-related decline in the percentage of diploid nuclei, coupled with an increase in the percentage of tetraploid and octaploid nuclei was observed in both groups. The major difference between the two groups was that the adult level of liver polyploidy was attained more slowly in the animals on dietary restriction as compared to the ad libitum fed controls. Polyploid cell formation in the liver is under the control of growth hormone, thyroid hormone and thymus, all of which might be influenced by dietary restriction.

Aging↗
PubMed1998

The titan mutants of Arabidopsis are disrupted in mitosis and cell cycle control during seed development.

We describe in this report a novel class of mutants that should facilitate the identification of genes required for progression through the mitotic cell cycle during seed development in angiosperms. Three non-allelic titan (ttn) mutants with related but distinct phenotypes are characterized. The common feature among these mutants is that endosperm nuclei become greatly enlarged and highly polyploid. The mutant embryo is composed of a few giant cells in ttn1, several small cells in ttn2, and produces a normal plant in ttn3. Condensed chromosomes arrested at prophase of mitosis are found in the free nuclear endosperm of ttn1 and ttn2 seeds. Large mitotic figures with excessive numbers of chromosomes are visible in ttn3 endosperm. The ttn1 mutation appears to disrupt cytoskeletal organization because endosperm nuclei fail to migrate to the chalazal end of the seed. How double fertilization leads to the establishment of distinct patterns of mitosis and cytokinesis in the embryo and endosperm is a central question in plant reproductive biology. Molecular isolation of TITAN genes should help to answer this question, as well as related issues concerning cell cycle regulation, chromosome movement and endosperm identity in angiosperms.

Arabidopsis↗
PubMed1976

Chromosomal characterisation of five lepidopteran cell lines of Malacosoma disstria (Lasiocampidae) and Christoneura fumiferana (Tortricidae).

Chromosome number and morphology have been examined in four established cell lines (Md63, Md66, Md108, and Md109) of the forest tent caterpillar, Malacosoma disstria Hübner, and one (Cf124) of the spruce budworm, Choristoneura fumiferana (Clemens). Chromosome number distributions of Md63 (mode = 112) Md108 (mode = 103), Md109 (mode= 103), and Cf124 (mode = 110) overlap sufficiently to prevent identification of individual lines by number alone. However, Md66 is exceptional in possessing a modal number of 157. One large chromosome occurs in cells of all lines. The presence of this chromosome, the lack of any distinct polyploid series among chromosome numbers encountered, and the general inverse relationship between number and size of chromosomes, suggest that the high level of heteroploidy characteristic of these and other lepidopteran cell lines reflects not only a possible polyploid origin but also extensive chromosomal rearrangement and fragmentation. Tolerance for such change is attributed to the holokinetic organisation of lepidopteran chromosomes. A distinct heteropycnotic body is present in about 10% of Cf124 cell nuclei, and can be used as a marker for this line. This body may represent the sex chromatin normally encountered in somatic cells of female C. fumiferana.

Cell Line↗
PubMed1998

Karyotypic evolution of cells in culture: a new concept.

The Chapter summarizes peculiarities of karyotypic variability during establishment and long-term cultivation of permanent cell lines. A new concept on pathways of karyotypic evolution of cells in culture is put forward. A detailed description is presented of the author's original approach of cytogenetic analysis of cell lines provided for a principally new characteristic of the cell line: its generalized reconstructed karyotype (GRK). Its use as a criterion to evaluate authenticity, purity, and stability of cell lines is discussed. Based on analysis of the GRK, two stages of karyotype evolution of cell lines are revealed: establishment and stabilization, different in karyotypic variability of the cell population and in peculiarities of clone selection. Comparison of peculiarities of karyotypic variability of leukemic and tumor cells both in vitro and in vivo was made, and general regularities of their karyotypic evolution have been established, such as nonrandom changes in the number and structure of chromosomes and deletion of one of the sex chromosomes, as well as regularities characteristic only of cells in culture in most human and animal cell lines (at least 85%) of disomy on all autosomes. The rest of the cell lines, 15%, are characterized by either partial or total monosomies on certain autosomes during long-term cultivation. Three main compensatory mechanisms of maintaining viability of cell lines that have lost genetic material are discussed: polyploidization of the initial cell clone, amplification of oncogenes (predominantly of mys family), and extracopying of whole autosomes or of their fragments.

Animals↗
PubMed1983

Characterization of a new human endometrial carcinoma (RL95-2) established in tissue culture.

A new human endometrial cell line, RL95-2, derived from a Grade 2 moderately differentiated adenosquamous carcinoma of the endometrium has been passaged successfully in cell culture for more than 2 yr. The cells are characteristically epithelioid with well-defined junctional complexes, tonofilaments, filopodialike extensions, and surface microvilli. Nuclei are large, irregular, and invaginated frequently with multiple, prominent, lamellar nucleoli. The cells have a log phase doubling time of 22 to 34 h followed by continued growth at a reduced rate with no apparent plateau phase. They exhibit a strong tendency for piling up as well as for the formation of glandlike dome structures. Karyotypically the line is trisomic 8 (47,XX,+8) and has an 8% frequency of polyploidization. Both cytoplasmic and nuclear estrogen receptors are present. Antihuman alpha-keratin characterizes the cell line as epithelial, nonstromal. The RL95-2 cell line may provide a useful in vitro system for the investigation of the endocrine regulation of endometrial neoplasia.

Adenocarcinoma↗
PubMed1996

The significance of elevated progesterone at the time of administration of human chorionic gonadotropin may be related to luteal support.

PURPOSE: Our purpose was to determine if the progesterone concentration on the day of HCG administration is associated with the establishment of pregnancy in IVF patients. METHODS: Concentrations of progesterone on the day of hCG were examined retrospectively in 293 patients with ovarian hyperstimulation induced by GnRH agonist with hMG and/or FSH. Patients were grouped based on progesterone concentration: < 0.9, 0.9 to < 1.2, 1.2 to < 1.5, and > or = 1.5 ng/ml. Oocytes recovered, fertilization rate, and pregnancy rate were compared among groups. RESULTS: Patients with a higher concentration of progesterone had increased concentrations of estradiol and greater numbers of oocytes retrieved. No differences were found for fertilization rate, polyploid fertilization, or pregnancy rate. It was noted that there was an association between the type of luteal support and the effect of high preovulatory progesterone on pregnancy rate. CONCLUSIONS: These data suggest that oocytes and embryos derived from patients with high preovulatory progesterone are not of a reduced quality. The association of high progesterone concentrations with a reduced rate of pregnancy varied with the type of luteal support.

Biomarkers↗
PubMed2001

Expression of scinderin in megakaryoblastic leukemia cells induces differentiation, maturation, and apoptosis with release of plateletlike particles and inhibits proliferation and tumorigenesis.

Rapid proliferation of atypical megakaryoblasts is a characteristic of megakaryoblastic leukemia. Cells from patients with this disorder and cell lines established from this type of leukemia showed the presence of gelsolin but the absence of scinderin expression, 2 filamentous actin-severing proteins present in normal megakaryocytes and platelets. Vector-mediated expression of scinderin in the megakaryoblastic cell line MEG-01 induced a decrease in both F-actin and gelsolin. This was accompanied by increased Rac2 expression and by activation of the PAK/MEKK.SEK/JNK/c-jun, c-fos transduction pathway. The Raf/MEK/ERK pathway was also activated in these cells. Transduction pathway activation was followed by cell differentiation, polyploidization, maturation, and apoptosis with release of platelet-like particles. Particles expressed surface CD41a antigen (glycoprotein IIb/IIIa or fibrinogen receptor), had dense bodies, high-affinity serotonin transport, and circular array of microtubules. Treatment of particles with thrombin induced serotonin release and aggregation that was blocked by CD41a antibodies. PAC-1 antibodies also blocked aggregation. Exposure of cells to PD98059, a blocker of MEK, inhibited antigen CD41a expression, increases in cell volume, and number of protoplasmic extensions. Cell proliferation and cell ability to form tumors in nude mice were also inhibited by the expression of scinderin. MEG-01 cells expressing scinderin had the same fate in vivo as in culture. Thus, when injected into nude mice, they entered apoptosis and released platelet-like particles. The lack of scinderin expression in megakaryoblastic leukemia cells seems to be responsible for their inability to enter into differentiation and maturation pathways characteristic of their normal counterparts.

Actins↗
PubMed2003

Characterization of a novel breast carcinoma xenograft and cell line derived from a BRCA1 germ-line mutation carrier.

A human tumor xenograft (L56Br-X1) was established from a breast cancer axillary lymph node metastasis of a 53-year-old woman with a BRCA1 germ-line nonsense mutation (1806C>T; Q563X), and a cell line (L56Br-C1) was subsequently derived from the xenograft. The xenograft carries only the mutant BRCA1 allele and expresses mutant BRCA1 mRNA but no BRCA1 protein as determined by immunoprecipitation or Western blotting. The primary tumor, lymph node metastasis, and xenograft were hypodiploid by DNA flow cytometry, whereas the cell line displayed an aneuploidy apparently developed via polyploidization. Cytogenetic analysis, spectral karyotyping, and comparative genomic hybridization of the cell line revealed a highly complex karyotype with numerous unbalanced translocations. The xenograft and cell line had retained a somatic TP53 missense mutation (S215I) originating from the primary tumors, as well as a lack of immunohistochemically detectable expression of steroid hormone receptors, epidermal growth factor receptor, human epidermal growth factor receptor 2 (HER-2), and keratin 8. Global gene expression analysis by cDNA microarrays supported a correlation between the expression profiles of the primary tumor, lymph node metastasis, xenograft, and cell line. We conclude that L56Br-X1 and L56Br-C1 are useful model systems for studies of the pathogenesis and new therapeutic modalities of BRCA1-induced human breast cancer.

Aneuploidy↗
PubMed1983

[Different modes of cell reproduction in the placental differentiation of mammals].

Summarized is the total evidence on the role of various modes of cell reproduction in the course of differentiation of various trophoblast population cells. The giant trophoblast cells, reaching a tremendously high (for mammals) degree of ploidy (512-4096c), blast cells, reaching a tremendously high for (mammals) degree of ploidy (512-4096c), give an example of cells that display a "non-classical" form of polyteny with not regular polyteny in chromosomes. For the trophoblast cells, a period of easy recognition with not regular, polyteny in chromosomes. For display a "non-classical form of polyteny with not regular polyteny in chromosomes. For the trophoblast cells, a period of easy recognition polytene chromosome has been established timed to definite stages of cell differentiation and of the polytene nucleus cycle. The start of phagocytic activity in trophoblast cells coincides in time with their transition to endoreduplication, both the processes terminating also simultaneously. It does not seem unlikely that such that each a correlation may appear due to a strict programmed appearance of numerous features of trophoblast cells, including their lifespan both in situ and under condition of -transplantation to different organs. Biological importance of various ways of reproduction in trophoblast cells (mitosis, non-completed polyploidizing mitosis, endoreduplication) in various parts of placenta is discussed in relation to their roles in development and functioning of this organo.

Animals↗
PubMedDate not supplied

Characterization of three cloned cell lines from a N-nitrosobis(2-hydroxypropyl)amine-induced transplantable hamster pancreatic ductal adenocarcinoma.

To investigate characteristics of pancreatic carcinoma growth behavior, the cloned cell lines, HPD1NR, HPD2NR, and HPD3NR, were established from a transplantable hamster ductal adenocarcinoma induced by N-nitrosobis(2-hydroxypropyl)amine (BHP). All three clones showed similar epithelial cell morphology and grew as sheets in culture with no differences in doubling times, ranging from 23-28 h. Mutation in the c-Ki-ras exon 1 was detected in common. The modal chromosome numbers were also found to be similar at 60, 62, and 60-62 in the less than tetraploid cells in the three clones. In contrast, a clear difference in frequencies of tetraploid or polyploid cells at 24.7, 22.5, and 75.5% in HPD1NR, HPD2NR, and HPD3NR, respectively, was evident. Tumorigenic potency evaluated by transplanting individual clones revealed HPD3NR to display pronouncedly less growth in syngeneic hamsters. The results suggest that increase in frequency of tetraploid or polyploid cells might be associated with a decreased in vivo growth potential of hamster pancreatic ductal adenocarcinomas, and suggest that these clones might become a valuable tool for understanding in vivo growth mechanisms of cancer cells.

Animals↗
PubMed1988

Characterization of a new megakaryocytic cell line: the Dami cell.

A new human megakaryocytic cell line (Dami) has been established from the blood of a patient with megakaryoblastic leukemia. The Dami cells grow primarily in suspension with a doubling time of 24 to 30 hours. By light and electron microscopy, the Dami cells range in size from 12 to 120 micron in diameter and have lobulated nuclei characteristic of megakaryocytes. At least 89% of the cells react with monoclonal antibodies against platelet glycoproteins (GP) Ib and IIB/IIIa, and glycophorin. The cells do not react with antibodies against lymphoid, monocyte, granulocyte, or macrophage antigens. Thirteen percent of the cells become polyploid, spontaneously achieving greater than 4N DNA ploidy levels. In response to phorbol myristate acetate (PMA), the proportion of cells with ploidy levels greater than 4N increased threefold and could be separated into discrete ploidy groups. PMA also increased the expression of GPIb, the GPIIb/GPIIIa complex,l and von Willebrand factor. Cytogenetic analysis revealed a human male hyperdiploid karyotype with a modal chromosome number of 54 to 64 and several consistent clonal chromosomal abnormalities. These included a partial deletion of chromosome 5 and a translocation involving chromosome 3. In contrast to other megakaryocytic cell lines in which only a small portion of the cells express the megakaryocytic phenotype, nearly all of the Dami cells express platelet glycoproteins. Thus, the Dami cells provide a superior model in which to study human megakaryocyte biochemistry and differentiation.

Antigens, Differentiation↗
PubMed1980

[Analysis of the depression of the mitotic activity of the corneal epithelium in white rats under chronic pyrogenal stress].

It was established previously that stable depression of cell division in corneal epithelium under chronic stress is not related to variations in the index of labeled nucleus (ILN). The paper demonstrates that five-day long pyrogenal injection did not alter the daily ILN: it was 67% in control rats and 69.7% in the experimental group. Cytophotometric logical sections showed that chronic stress led to a significant increase in the number of tetraploid nuclei (from 3.2 to 7.7%). No material changes were observed in the content of nuclei belonging to other classes. No polyploidization or G(2) growth evidence were recorded either. The absence of progressive rise in tetraploid nuclei and of more pronounced polyploidization in the presence of long-term depression of the cell cycle and stable ILN is explained by circadian variations in the mitotic activity reaction to stress. During chronic stress the depression of the cell cycle observed in the day and evening time was not recorded in the morning hours at the height of the mitotic activity. This circadian shunt is conducive to maintaining modal ploidity of corneal nuclei.

Animals↗