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PubMed1999

Mechanisms of genetic instability revealed by analysis of yeast spindle pole body duplication.

Aneuploidy and polyploidy are commonly observed in transformed cells. These states arise from failures during mitotic chromosome segregation, some of which can be traced to defects in the function or duplication of the centrosome. The centrosome is the organizing center for the mitotic spindle, and the equivalent organelle in the budding yeast, Saccharomyces cerevisiae, is the spindle pole body. We review how defects in spindle pole body duplication or function lead to genetic instability in yeast. There are several well documented instances of genetic instability in yeast that can be traced to the spindle pole body, all of which serve as models for genetic instability in transformed cells.

Aneuploidy↗

PubMed2000

[Chromosomal variability of ginseng cells transformed with plant oncogene rolC].

Chromosome numbers were was studied in ginseng cell line 1c transformed with Agrobacterium rhizogenes strain A4, which carried plasmid pRiA4, and with A. tumefaciens strain GV3101, which carried vector pPCV002-35S rolC. As compared with the nontransformed cell line 1c, tumor cell cultures 1c-A4 and 1c-rolC and the tissues of rolC teratoma (excluding leaves) displayed higher polyploidy and aneuploidy. The 1c-A4 and 1c-rolC hairy-root cultures also had aneuploid and polyploid cells, but the chromosome variation was lower than in tumor cells or the initial culture 1c. Generally, an increase of chromosome variation in cultivated cells was the main effect of the integration of several oncogenes, which were in the A. rhizogenes A4 T-DNA, or of the individual rolC gene in the ginseng genome. Another effect consisted in stabilization of the chromosome number in some differentiated transgenic tissues. Possible reasons for this effect are discussed.

Cell Line, Transformed↗

PubMed2000

Geminiviruses: models for plant DNA replication, transcription, and cell cycle regulation.

Geminiviruses have small, single-stranded DNA genomes that replicate through double-stranded intermediates in the nuclei of infected plant cells. Viral double-stranded DNA also assembles into minichromosomes and is transcribed in infected cells. Geminiviruses encode only a few proteins for their replication and transcription and rely on host enzymes for these processes. However, most plant cells, which have exited the cell cycle and undergone differentiation, do not contain the replicative enzymes necessary for viral DNA synthesis. To overcome this barrier, geminiviruses induce the accumulation of DNA replication machinery in mature plant cells, most likely by modifying cell cycle and transcriptional controls. In animals, several DNA viruses depend on host replication and transcription machinery and can alter their hosts to create an environment that facilitates efficient viral replication. Analysis of these viruses and their proteins has contributed significantly to our understanding of DNA replication, transcription, and cell cycle regulation in mammalian cells. Geminiviruses have the same potential for plant systems. Plants offer many advantages for these types of studies, including ease of transformation, well-defined cell populations and developmental programs, and greater tolerance of cell cycle perturbation and polyploidy. Our knowledge of the molecular and cellular events that mediate geminivirus infection has increased significantly during recent years. The goal of this review is to summarize recent research addressing geminivirus DNA replication and its integration with transcriptional and cell cycle regulatory processes.

Cell Cycle↗

PubMed2000

The role of p53 in the response to mitotic spindle damage.

The p53 tumour suppressor protein has defined roles in G1/S and G2/M cell cycle checkpoints in response to a range of cellular stresses including DNA damage, dominant oncogene expression, hypoxia, metabolic changes and viral infection. In addition to these responses, p53 can also be activated when damage occurs to the mitotic spindle. Initially, spindle damage activates a p53-independent checkpoint which functions at the metaphase-anaphase transition and prevents cells from progressing through mitosis until the completion of spindle formation. Cells eventually escape from this block (a process termed 'mitotic slippage'), and an aberrant mitosis ensues in which sister chromatids fail to segregate properly. After a delay period, p53 responds to this mitotic failure by instituting a G1-like growth arrest, with an intact nucleus containing 4N DNA, but without the cells undergoing division. Cells lacking wild-type p53 are still able to arrest transiently at mitosis, and also fail to undergo division, underscoring that the delay in mitosis is p53-independent. However, these cells are not prevented from re-entering the cell cycle and can reduplicate their DNA unchecked, leading to polyploidy. Additionally, p53-null cells which experience spindle failure often show the appearance of micronuclei arising from poorly segregated chromosomes which have decondensed and been enclosed in a nuclear envelope. The ability of p53 to prevent their formation suggests an additional G2 involvement which prevents nuclear breakdown prior to mitosis. The molecular mechanism by which p53 is able to sense mitotic failure is still unknown, but may be linked to the ability of p53 to regulate duplication of the centrosome, the organelle which nucleates spindle formation.

Animals↗

PubMed2000

PKC412--a protein kinase inhibitor with a broad therapeutic potential.

The staurosporine derivative PKC412 was originally identified as an inhibitor of protein kinase C (PKC) and subsequently shown to inhibit other kinases including the kinase insert domain receptor (KDR) (vascular endothelial growth factor receptor, VEGF-R2), the receptor of platelet-derived growth factor, and the receptor for the stem cell factor, c-kit. PKC412 showed a broad antiproliferative activity against various tumor and normal cell lines in vitro, and was able to reverse the Pgp-mediated multidrug resistance of tumor cells in vitro. Exposure of cells to PKC412 resulted in a dose-dependent increase in the G2/M phase of the cell cycle concomitant with increased polyploidy, apoptosis and enhanced sensitivity to ionizing radiation. PKC412 displayed a potent antitumor activity as single agent and was able to potentiate the antitumor activity of some of the clinically used cytotoxins (Taxol and doxorubicin) in vivo. The combined treatment of PKC412 with loco-regional ionizing irradiation showed significant antitumor activity against tumors which are resistant to both ionizing radiation and chemotherapeutic agents (dysfunctional p53). The finding that PKC412 is an inhibitor of the VEGF-mediated cellular signaling via inhibition of KDR and PKC in vitro is consistent with the in vivo inhibition of VEGF-dependent angiogenesis in a growth factor implant model. Orally administered PKC412 also strongly inhibited retinal neovascularization as well as laser-induced choroidal neovascularization in murine models. In summary, PKC412 may suppress tumor growth by inhibiting tumor angiogenesis in addition to directly-inhibiting tumor cell proliferation via its effects on PKC and/or other protein kinases. PKC412 is currently in Phase I clinical trials for treatment of advanced cancer as well as for the treatment of ischemic retinopathy.

Angiogenesis Inhibitors↗

PubMed2000

Comparison of 2-methoxyestradiol-induced, docetaxel-induced, and paclitaxel-induced apoptosis in hepatoma cells and its correlation with reactive oxygen species.

BACKGROUND: Previously, the authors observed that paclitaxel treatment of hepatoma cells resulted in differential cytotoxicity. Whether other antimicrotubule agents (docetaxel and 2-methoxyestradiol) are more effective than paclitaxel is not clear. Moreover, whether the modulation of reactive oxygen species (ROS) is involved in the drug-induced growth inhibition of hepatoma cells is not known. METHODS: The authors examined the effects of 2-methoxyestradiol, paclitaxel, and docetaxel on HepG2, Hep3B, HA22T/VGH, and Hepa1-6 hepatoma cell lines. The parameters examined included cell viability, cell membrane permeability, cell cycle distribution, DNA fragmentation, and ROS generation. RESULTS: Docetaxel and paclitaxel inhibited the growth of hepatoma cells at submicromolar concentrations, whereas that of 2-methoxyestradiol was within a micromolar range. This drug-induced growth inhibition was cell cycle dependent. 2-Methoxyestradiol-treated (10-50 microM) cells resulted in G2/M block prior to apoptosis. High dose (0.1 microM) docetaxel- and paclitaxel-treated cells resulted in a G2/M arrest followed by generation of polyploidy or apoptosis; however, low dose (0.01 microM) treatment induced apoptosis without G2/M arrest. The low dose effect was more significant in docetaxel-treated cells than in paclitaxel-treated cells. Although these antimicrotubule agents increased the formation of ROS, antioxidant treatment did not block drug-induced cell cycle and growth inhibition effects. CONCLUSIONS: The current results suggest that the growth inhibition of hepatoma cells induced by 2-methoxyestradiol, paclitaxel, and docetaxel was mediated through G2/M-phase arrest, caspase activation, and DNA fragmentation. The drug-induced apoptosis was independent of ROS formation. Docetaxel was more effective than paclitaxel in killing hepatoma cells. The potential of using 2-methoxyestradiol and docetaxel for the treatment of patients with hepatoma is worthy of further study.

2-Methoxyestradiol↗

PubMed2000

Intratumoral regional differences of DNA ploidy of gastrointestinal stromal tumors: a flow cytometric study.

BACKGROUND: Mesenchymal neoplasms of the gastrointestinal tract remain controversial in regard to both diagnostic criteria and prognostic factors. In order to investigate whether DNA ploidy in a single area of a tumor can be representative of the entire tumor, multiple samples from the same tumor were analyzed in 79 cases of gastrointestinal stromal tumors. METHODS: Forty-three male and 36 female patients, aged 11 to 80 years with stromal tumors of the GI tract were selected. Flow cytometric DNA ploidy analysis was done on tissue cut from paraffin blocks by using the modified technique of Hedley et al. RESULT: All 34 benign stromal tumors displayed a concordant diploid DNA content. Eighteen (40%) of the 45 malignant stromal tumors (13 low-grade, 5 high-grade) were diploid. Aneuploid and polyploid DNA stemlines were found in 10 cases (43.4%) of low-grade stromal tumors, and in 17 cases (77.2%) of high-grade stromal tumors. DNA polyploidy was identified in 21 (46.7%) of 45 malignant stromal tumors. Also, aneuploid populations were accompanied by diploid populations. CONCLUSION: DNA content correlated well with histologic category. Tumor aneuploidy was more frequently seen in tumors of advanced stage, but no relationship was seen between tumor stage and the presence of tumor heterogeneity. Differences in ploidy levels were thus not necessarily associated with apparent differences in histologic patterns. The mechanism responsible for regional differences in DNA ploidy in the same tumor remain unexplained. Several hypotheses are reviewed.

Adolescent↗

PubMed2000

Allozyme evidence for genetic autopolyploidy and high genetic diversity in tetraploid cranberry, Vaccinium oxycoccos (Ericaceae).

Polyploidy has been important in the evolution of angiosperms and may significantly affect population genetic diversity and structure. Nineteen isoenzyme loci were studied in diploid and tetraploid populations of Vaccinium oxycoccos (Ericaceae), and the results are compared with data previously reported for the related V. macrocarpon. Diploid V. oxycoccos and V. macrocarpon were readily discriminated based on their allozymic variation. No evidence for fixed heterozygosity was found in tetraploid V. oxycoccos. In contrast, all polymorphic loci exhibited both balanced and unbalanced heterozygotes, with some individuals exhibiting a pattern consistent with the presence of three alleles. These results support an autopolyploid origin for tetraploid V. oxycoccos. However, tetraploid V. oxycoccos possessed a suite of alleles not found in diploid V. oxycoccos; half of these alleles were shared with V. macrocarpon. This suggests that autotetraploid V. oxycoccos may have undergone hybridization with V. macrocarpon or that the autotetraploid retained the genetic variation present in an ancestral diploid species. Following theoretical expectations, proportion of polymorphic loci, mean number of alleles, and observed heterozygosity were significantly higher for the autotetraploid than for the diploid. Mean inbreeding (F(IS)) was similar for diploid and tetraploid V. oxycoccos. The latter exhibited population differentiation (F(ST)) exceeding both diploid species.

Journal Article↗

PubMed2000

Origin and relationships of the tarweed-silversword lineage (Compositae-Madiinae).

Based on results from phylogenetic analyses of nuclear 18S-26S rDNA internal transcribed spacer (ITS) region sequences, we suggest that the monophyletic tarweed and silversword subtribe (Madiinae) is phylogenetically nested among epaleate, x = 19 species of helenioid Heliantheae. Strong bootstrap support (100%) was obtained for a sister-group relationship between Madiinae and Arnica (including Mallotopus and Whitneya) in an analysis including representatives of recognized genera in a principally Californian clade (Madieae sensu Baldwin) identified from a phylogenetic investigation of Heliantheae s.l. (sensu lato) and Eupatorieae. In all minimum-length trees, the robust lineage comprising Madiinae and Arnica (x = 19) is part of a larger clade that also comprises Eatonella s.s. (sensu stricto), Hulsea, and Venegasia, all with x = 19. The phylogenetic position of Madiinae within a group of genera based uniformly on x = 19 leads us to conclude that the modal numbers of n = 7 and n = 8 (and other numbers, as low as n = 4) in Madiinae are the results of extreme dysploidy. Among the x = 19 "arnicoid" taxa, the near-universal characteristics of perenniality (except in the monotypic Eatonella s.s. and a minority of hulseas) and montane or high-latitudinal occurrence (except in the monotypic Venegasia) lead us to suggest that the most recent common ancestor of the tarweeds (a principally annual group of seasonally dry, low-elevation habitats) was probably a montane, herbaceous perennial resembling the unusual subalpine and alpine tarweeds constituting Raillardella s.s. (x = 17), an arnica-like genus. In Madiinae, Raillardella s.s. may be plesiomorphic in habit, capitular and ecological characteristics, and high base chromosome number. Shifts to an annual habit and to low chromosome numbers in Madiinae have been followed by subsequent episodes of polyploidy and descending dysploidy. We conclude that genome evolution in Madiinae has been marked by wide swings in chromosome number that confuse identification of diploids and polyploids.

Journal Article↗

PubMed2001

TP53-dependent chromosome instability is associated with transient reductions in telomere length in immortal telomerase-positive cell lines.

Telomere shortening in telomerase-negative somatic cells leads to the activation of the TP53 protein and the elimination of potentially unstable cells. We examined the effect of TP53 gene expression on both telomere metabolism and chromosome stability in immortal, telomerase-positive cell lines. Telomere length, telomerase activity, and chromosome instability were measured in multiple clones isolated from three related human B-lymphoblast cell lines that vary in TP53 expression; TK6 cells express wild-type TP53, WTK1 cells overexpress a mutant form of TP53, and NH32 cells express no TP53 protein. Clonal variations in both telomere length and chromosome stability were observed, and shorter telomeres were associated with higher levels of chromosome instability. The shortest telomeres were found in WTK1- and NH32-derived cells, and these cells had 5- to 10-fold higher levels of chromosome instability. The primary marker of instability was the presence of dicentric chromosomes. Aneuploidy and other stable chromosome alterations were also found in clones showing high levels of dicentrics. Polyploidy was found only in WTK1-derived cells. Both telomere length and chromosome instability fluctuated in the different cell populations with time in culture, presumably as unstable cells and cells with short telomeres were eliminated from the growing population. Our results suggest that transient reductions in telomere lengths may be common in immortal cell lines and that these alterations in telomere metabolism can have a profound effect on chromosome stability.

Cell Line, Transformed↗

PubMed2001

Comparative characterization of cell death between Sf9 insect cells and hybridoma cultures.

Physiological cell death (PCD) in Sf9 insect cell batch cultures was comprehensively characterized using simultaneous determinations of qualitative and quantitative assays, including agarose gel electrophoresis, confocal, epifluorescence, and transmission electron microscopy, and DNA content by flow cytometry. Results were compared to hybridoma cultures where abundant information of apoptosis exists. Both cultures shared some typical apoptosis features, including cell shrinkage, loss of sphericity, swollen endoplasmic reticulum and Golgi apparatus, chromatin condensation, and specific DNA degradation. However, distinctive morphological and kinetic differences between both cultures revealed that Sf9 cells died by an atypical PCD process characterized by absence of nuclear fragmentation, scarce association of condensed chromatin to the nuclear envelope, swollen mitochondria, and high nonspecific DNA degradation. These features, distinctive of necrosis, were not observed in the normal apoptotic process of hybridomas. Glucose depletion marked the appearance of apoptotic Sf9 cells, which there up on increased gradually, whereas apoptotic hybridomas rapidly increased upon glutamine depletion. Furthermore, active phagocytosis was found in Sf9 viable cells, a characteristic phenomenon during in vivo apoptosis but uncommon for in vitro cultures. Sf9 cells contained unusually high numbers of phagosomes, particularly after glucose depletion. Additionally, few apoptotic bodies accumulated in culture, suggesting their elimination by phagocytosis. Other distinctive characteristics of Sf9 cells were the presence of a polynucleated hypertrophic population fraction, polyploidy, cell cycle arrest in G2/M phase, and more necrosis compared to hybridomas. Such phenomena prevented a reliable quantification of apoptosis from determination of the sub-G1 peak. Nonetheless, emergence of a bimodal Sf9 cell size distribution coincided with the increase in the sub-G1 population and onset of death. The fraction of particles in the smaller peak (6-11 microm diameter) closely correlated with the fractions of apoptotic bodies, late apoptotic, and secondary necrotic cells. Accordingly, Sf9 cell size was shown to be an effective, rapid, and simple parameter for quantifying death. Altogether, the results of this study provide new insights into PCD and other phenomena in insect cell culture important for biotechnological applications of Sf9 cells.

Aneuploidy↗

PubMed2001

Barriers to interbreeding in the Eriophyllum lanatum (Asteraceae, Helenieae) species complex.

Sampling 426 plants from 271 populations of Eriophyllum lanatum, a western North American species complex, revealed 195 diploid (x = 8), 61 tetraploid, ten hexaploid, and five octoploid populations. Polyploids were ∼400% more frequent in taxonomic intermediates. One to four supernumerary chromosomes were found in 13% of the populations. Artificial hybridizations were made in 58 of the 65 possible diploid-level combinations involving the ten varieties of E. lanatum and E. confertiflorum var. confertiflorum. Aberrations, mostly failure to pair normally, were observed in diakinesis or M(1) cells of progeny in 23 of 99 crosses. Studies of pollen stainability in cotton blue-lactophenol and other fertility indicators in 886 F(1)'s from 191 crosses involving 81 populations showed that strong (22-40% pollen stainability) to weak (60-76% pollen stainability) barriers to interbreeding existed among diploids of the E. lanatum varieties and among them and E. confertiflorum var. confertiflorum. Pollen stainability was much higher in progenies of tetraploid, hexaploid, and octoploid intra- and interspecific crosses involving E. confertiflorum var. tanacetiflorum, E. jepsonii, and E. latilobum than in diploid ones, supporting the hypothesis that polyploidy has mainly served to stabilize the products of intervarietal and interspecific hybridizations.

Journal Article↗

PubMed2001

Genomic and genetic relationships among species of Leymus (Poaceae: Triticeae) inferred from 18S-26S ribosomal genes.

The 18S-26S ribosomal genes in three closely related species of Leymus (Poaceae: Triticeae) were examined using fluorescence in situ hybridization (FISH) and restriction fragment length polymorphism (RFLP). Both approaches revealed a close relationship between L. arenarius (8x = 56, northern European) and L. racemosus (4x = 28, central Eurasian), whereas L. mollis (4x = 28, northern American/Pacific) was distinct. Each species had three homologous pairs of major rDNA loci: a1, a2, and a3 for L. arenarius; m1, m2, and m3 for L. mollis; and r1, r2, and r3 for L. racemosus. Leymus arenarius had in addition three minor loci, a4, a5, and a6. The major loci of L. arenarius and L. racemosus were identical, indicating that the former species could have originated from the latter, via interspecific hybridization and/or polyploidy. The rDNA-RFLPs further indicated relationships of these species to other species of Leymus (L. karellini, 8x = 56 and L. angustus, 12x = 84) and Psathyrostachys (P. fragilis, P. huashanica, P. juncea, and P. lanuginosa, which are all diploids). A phenogram constructed from 20 BamHI, EcoRI, and DraI rDNA fragments revealed closer relationship between the two genera, Leymus and Psathyrostachys, than that among species within a genus.

Journal Article↗

PubMed2001

Karyotype analysis in Brachiaria (Poaceae) species.

This is the first karyotype characterization of Brachiaria species. Twelve accessions belonging to five species were analysed. The basic chromosome number was x = 9 and 7, the same reported for the tribe Paniceae. Variations in the chromosome number were observed in B. decumbens (2n = 18; 36) and B. humidicola (2n = 36; 42; 54). Chromosome numbers of 2n = 18 in B. ruziziensis and 2n = 36 in B. brizantha and B. jubata were recorded. Inter- and intraspecific karyotype differentiation of the accessions analysed was facilitated by variations in karyotypic symmetry. The karyotypes were generally considered symmetrical, with a tendency to asymmetry in the direction of the polyploids. It is suggested that addition, deletions and mainly polyploidy have been the most direct causes involved in the chromosome evolution of this genus.

Chromosomes↗

PubMed2001

[Cytological aspects of similarity and difference of Myxozoa and Cnidaria].

A comparative cytomorphological analysis of Myxozoa and parasitic Cnidaria Polypodium hydriforme has been carried out in view of the Weill (1938) hypothesis, which regards Myxozoa as a reduced Cnidaria. The question on the relation of Myxozoa and Cnidaria was arising several times with the application of some new methods during the Myxozoa studies. At present the idea on their phylogenetic relationships has appeared again in connection with an absolutely new understanding of the myxozoan life cycle (Wolf, Markiw, 1984), as well as with the application of molecular-biological methods for their phylogenetic studies. The latter, however, provided some diverse results. So far no comparative cytomorphological analysis of Myxozoa and Polypodium has been carried out. The present paper is to fill the gap on the basis of accumulated facts. According to Weill (1938), the features of similarity of parasitic Cnidaria and Myxozoa are the following: 1) the presence in both of extrusomes (nematocysts and polar capsules) whose structure and development are surprizingly similar; 2) the nuclear dimorphism and somato-generative segregation; 3) the presence of a somatic nutritional cell, surrounding the multiplying generative cells; at present it is known that polyploidy of somatic nuclei and the absence of parasitophorous vacuole are characteristic of trophamnion of Polypodium and trophozoite of Myxozoa; 4) the presence of radial symmetry in both groups; 5) the construction of a diblastic organism made of a cluster of endodermal cells and a few ectodermal cells; 6) the similarity of their cell contacts (Grassé, 1970). At present it is possible to add to Weill's (1938) list of features common for parasitic Cnidaria and Myxozoa the number of important similarities between Polypodium and Myxozoa, some of which being not characteristic of Cnidaria: 1) the "cell in cell" organization of all Polypodium parasitic stages and all Myxozoa life cycle stages; 2) the presence of gametophore supplied with extrusomes; 3) both organisms have haplophase in their life cycles preceded by two-step meiosis; 4) there are mitochondria with tubular cristae in both organisms; 5) the absence of spermatozoa and eggs in both organisms; 6) the similarity of Polypodium cnidocile structure and the cone-like formation situated at the anterior end of polar capsule of actinospore (Lom. Dykova, 1997); 7) the participation of MTOC in the formation of extrusomes in both animals. In spite of the obvious similarity between Myxozoa and parasitic Cnidaria (including Polypodium) it is, however, necessary to take into account differences between them, the main being as follows: the absence in Myxozoa of flagellated stages, centrioles, tissues and organs, true blastophylla, planula-like larvae, gastrulation; the presence of low cell integrations in Myxozoa; Cnidaria and Myxozoa have different types of mitosis, their life cycles and the discharge mechanism of their stinging apparatus being also different. We consider as quite valid a suggestion by Siddall et al. (1995) that parasitic Cnidaria could present an early separated branch of the cnidarian evolution. Further studies of Myxozoa life cycle may show their more definite relation to parasitic Cnidaria. The problem has not yet been solved completely since the available molecular-biological data are rather contradictory and moreover there is no distinct idea as to the Eumetazoa ancestor so far. A further thorough investigation is badly needed in the feelds of developmental cycle, cytomorphology and molecular biology of the variety of narcomedusae and representatives of Myxozoa. This may help to find some transitional forms and stages of the animals and to understand whether we deal with a regressive evolution of parasitic Cnidaria or with a parallel evolution of taxa originated from the common ancestor.

Animals↗

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↗

PubMed2001

Quantitative assessment of oxyphilic cell lesions of the thyroid gland on fine needle aspiration samples.

OBJECTIVE: To assess the possible contribution by a multiparametric quantitative approach to the cytologic diagnosis of oxyphilic cell (OC) thyroid lesions. STUDY DESIGN: Ten cases of chronic lymphocytic (Hashimoto) thyroiditis and 10 nodular goiters containing oxyphilic cells plus 20 cases of tumors subsequently classified as oxyphilic cell adenomas (10 cases) or oxyphilic cell well-differentiated carcinomas (10 cases) were evaluated. The study was performed on May-Grünwald-Giemsa-stained smears for planimetric measurements. The same smears were destained and Feulgen restained for densitometric measurements. The latter were performed using static cytometry equipment measuring 100 and 20-30 lymphocytes per case for the determination of integrated optical density (IOD). The following parameters were considered: nuclear area, perimeter, maximum diameter, form ELL, form PE, IOD, 5c exceeding rate (5cER) and visual classification of histograms as euploid, polyploid and aneuploid. RESULTS: Mean nuclear area of carcinomas was smaller than that of adenomas, goiter and thyroiditis. Nuclear area was larger in adenomas than in other benign lesions and carcinomas. All the other planimetric parameters were similar in the lesions examined. Four carcinomas and three adenomas were aneuploid, and all the rest were euploid. All the cases of thyroiditis and goiter were euploid or polyploid; four thyroiditis cases showed polyploid histograms and 5cER values > 1. CONCLUSION: Morphometric and densitometric procedures have a limited role in the discrimination of OC lesions, but small nuclear area values may be useful in distinguishing OC carcinoma from other lesions. The role of densitometry seems even more limited because aneuploid histograms may be found among adenomas and carcinomas. Further studies are needed to explain polyploidy and 5cER > 1 in Hashimoto thyroiditis.

Biopsy, Needle↗

PubMedDate not supplied

[Deletion of YNZ22 and ALU-VPA/MYCL1 loci in human colonic adenocarcinoma and postoperative prognosis].

Since deletions of the short arm of chromosome 17 are the most common genetic defects in human colorectal carcinoma (CC), we tested the YNZ22 locus (D17S30, 17p13.3) for loss of heterozygosity (LH) in adenocarcinoma and in the normal colonic mucosa of 49 CC patients, and studied the association of LH with clinicomorphological features of the tumor. Allele frequency distribution of YNZ22 did not differ for the patients and healthy people. LH in YNZ22 in the tumor was found in 33% (13/39) of all informative cases, its frequency being thrice higher in men than in women (chi 2 = 5.21, p = 0.022). The defect was associated with moderate or poor histological differentiation (P2 = 0.0055) and polyploidy > 3n (P2 = 0.0035) of tumor cells and with high incidence of post-surgery relapse or metastasis. Analysis of both YNZ22 and Alu-VpA/MycL1 (1p34.3) loci in the tumor allowed reliable relapse prognosis in 76% of the CC patients. The probability of post-surgery relapse or metastasis was estimated at no less than 67% for patients with LH in at least one of the two loci in the tumor, and at somewhat more than 20% for patients without LH.

Adenocarcinoma↗