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A P Dyban

Publications and source records attributed to A P Dyban.

At least 19 recordsLinked to original sources
PubMed1999

Persistence of human mitochondrial DNA throughout the development to the blastocyst of mouse zygotes microinjected with human mitochondria.

The conditions for transfer of human mitochondria into fertilised mouse ova were elaborated. Species-specific primers were designed to discriminate human mitochondrial DNA (mtDNA) and the endogenous mtDNA in the preimplantation embryos. Human mitochondria isolated from the HepG2 cell line were microinjected into murine zygotes, and the latter cultured for 96 h to the blastocyst stage. The polymerase chain reaction allowed the detection of human mtDNA at every stage of embryo cleavage. In some cases a clear disparity in distribution of human mtDNA among blastomeres was evident.

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PubMed1997

The behaviour of mitochondria and cell integration during somatic hybridisation of sister blastomeres of the 2-cell mouse embryo.

The capacity of sister blastomeres of mouse embryos for induced fusion changed during the 2-cell stage. It was at low level (24%) at the early 2-cell stage, increased and reached 98.5% at the middle 2-cell stage and fell sharply to 31% at the late 2-cell stage. At the time corresponding to the G2/M-phase of the cell cycle the blastomeres fused in only 8% of cases. Vital staining of 2-cell embryos by rhodamine 123 showed that the mitochondria were dispersed throughout the cytoplasm with a ring-like (around the nucleus) or spot-like (over the metaphase plate) concentration in the centre of each blastomere. At the periphery of blastomeres the mitochondrial content was low. The behaviour of the mitochondria reflected the subsequent events of structural and functional integration of the sister blastomeres under induced fusion: a discernible boundary between partners during 30 min after electrofusion or 1 h after fusion with polyethylene glycol; movement of the two 'rings' to the centre of the blastomere fusion products (BFP) to form one large bright 'spot' over the common metaphase plate; mitochondria outlining the shape of the spindle and connection between sister blastomeres until completion of the first mitosis of BFP. The data obtained suggest that fusion of the blastomeres does not lead to extensive changes in the hybrid cytoplasm and integration of nuclear material is taking place only at metaphase stage. Cytogenetic examination of BFP at the 2-cell stage confirmed reconstruction of the tetraploid embryos and found that sister blastomeres of such embryos could asynchronously enter the next cleavage division similarly to normal diploid 2-cell embryos.

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PubMed1994

[The quantitative study of the areas of active rDNA location and of the other parameters of the interphase nucleolus detectable by silver staining during the cell differentiation of the rat trophoblast].

Different quantitative parameters of nucleolar silver staining have been studied in the cambial rat trophoblast cells on the 12th, 13th and 14th days of gestation. It has been shown that the number of Ag-positive granules in the nucleoli varied from 10 to 120. The number and the total area of silver stained granules in the nuclei increased progressively in the course of polyploidization, but was not doubled passing to the next ploidy level. Nevertheless, nucleolar area increased proportionally to the ploidy degree. The mean number and the total area of Ag-stained granules as well as the nucleolar area estimated for each ploidy level did not change significantly in the course of placenta development, suggesting an unchanged level of NOR activity at the studied stages of trophoblast cell differentiation. The data obtained on the interphase nucleoli differ from the data of the analysis of the metaphase Ag-NOR at the same period of placenta development, suggesting a diversity in the interphase and metaphase NOR organization. A proportion of cells with different number of nucleoli in the cambial rat trophoblast cells was maintained unchanged in the studied period of the placenta development, the majority (80-90%) of cells contained from 1 to 3 nucleoli. Such a proportion was similar in the cells of different levels of ploidy up to 16c. In this connection the association of NORs is suggested to be in relation with switching from the polyploid mitotic cycle to the endoreduplication leading to polyteny.

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PubMed1993

Okadaic acid induces premature chromosome condensation reflecting the cell cycle progression in one-cell stage mouse embryos.

Haploid parthenogenetic embryos as well as fertilized mouse eggs were treated in vitro with 1-10 microM okadaic acid (OA) at the one-cell stage. Cytogenetic analysis detected that OA induces nuclear envelope breakdown (NEBD) and premature condensation of interphase chromosomes in pronuclei as well as in 2nd polar body (PB) nuclei. G1-, S-, and G2-type prematurely condensed chromosomes (PCC) were found in pronuclei of embryos of different age, which reflects their progression through the first cell cycle. In nuclei from 2nd PBs only G1- and S-type PCC were observed. Using the types of PCC as a criterion of different phases of the cell cycle, it was possible to estimate that in haploid parthenogenetic embryos G1-phase lasts until 5.5 hr post activation (hpa), S-phase takes from 4.5 to 9.5 hpa, and from 8.5 hpa G2-phase had started. Second PBs were found to be in G1-phase until 6.5 hpa and S-phase started in some as early as 5.5 hpa, but in most not before 7.5 hpa. Treatment with OA visualizes G1-chromosomes in pronuclei as well as in 2nd PBs, and it is easy to count the number of these chromosomes and recognize a T6 marker chromosome. The possibility to apply cytogenetic analysis of G1-chromosomes from 2nd PBs for a more accurate detection of maternal meiotic nondisjunction is discussed.

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PubMed1993

[The frequency of sister chromatid exchanges in preimplantation mouse embryos].

The frequency of sister-chromatid exchanges (SCE) was analysed in spontaneously and superovulated morulae and blastocyst of CBA, C57Bl, F1 CBA x C57Bl mice in culture with bromodeoxyuridine (BrdU). The background level for SCE was found to be 4-5 times higher in early embryos than in fetal or adult tissues of the mouse. The phenomenon of spontaneous sister-chromatid differentiation (SCD) of blastocyst chromosomes without any treatment with BrdU was observed. The SCE frequency was calculated on such chromosomes and was the same as with BrdU. A possible mechanism of this phenomenon is suggested.

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PubMed1992

Visualization of second polar body chromosomes in fertilized and artificially activated mouse oocytes treated with okadaic acid.

OBJECTIVE: Our objective was to develop a new reliable method for cytogenetic analysis of the chromosome set in second polar bodies (PBs) from one-cell-stage mouse embryos. SETTING: The study took place at the Reproductive Biology and Experimental Cytogenetics Laboratories. METHODS: Oocytes from F1 hybrid and T6/T6 mice were fertilized in vitro and artificially activated with ethanol. Zygotes, parthenogenetic embryos, and isolated second PBs were treated with 10 microM okadaic acid (OA) for 1-2 hr, further cultured in plain medium, and fixed. Chromosomal preparations were made and C-banded, and the number of chromosomes in second PBs and embryos was counted. RESULTS: OA-induced nuclear envelope breakdown in pronuclei as well as in second PB nuclei. Countable chromosome plates were obtained in 92-93% of second PBs treated 4-4.5 hr after activation. The T6 marker chromosome could easily be recognized in second PBs from T6/T6 mice. A haploid set of chromosomes was obtained in 18 of 19 isolated second PBs treated with OA 4-5 hr after activation. CONCLUSION: Treatment of second PBs with OA allows visualization of the PB chromosomes. Cytogenetic analysis of the second PB and the corresponding oocyte constitutes a new approach for the study of meiotic nondisjunction in experimental cytogenetics. The chromosomal study of isolated second PBs seems to be promising for clinical preimplantation cytogenetics.

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PubMed1991

[The nucleolus organizer regions of the metaphase chromosomes in cambial cell populations of mouse and rat placenta].

The number of silver-stained nucleolus organizing regions (Ag-NORs) was counted in metaphase plates of the fetal part of placenta of mice and rats and in the tissues of their embryos. On day 9 of mouse gestation and on day 12 of rat gestation, up to 82% of metaphases in the fetal part of placenta have the highest possible number of chromosomes with Ag-NORs (9-10 for mice and 5-6 for rats). In the later embryogenesis (day 10 for mice and day 14 for rats), a great number of metaphases have either no Ag-NORs (34.9% for mice, 17.9% for rats), or only 1-2 Ag-NOR-chromosomes (17.4% for mice and 14.5% for rats). But in the tissues of embryos being on the same embryonic stages the changes in frequency of metaphases with different numbers of Ag-NOR-chromosomes have proved to be less obvious. In the differentiated polyploid placenta cells the number of chromosomes with Ag-NORs can be considerably lower than in diploid ones. It is proposed that the changes in frequency of metaphases with different numbers of Ag-NOR-chromosomes may reflect the regulation of ribosomal gene transcription in the embryogenesis, in particular, in differentiation of cambial cell populations of rodent placenta.

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PubMed1990

The silver-stained NOR and argentophilic nuclear proteins in early mouse embryogenesis: a cytological study.

Silver staining (Howell and Black, 1980) was used in light and electron microscopic studies for detecting the localization of argentophilic nuclear proteins in fertilized ova and cleaving mouse embryos. No silver-stained nucleolus organizing regions (NORs) (Ag-NORs) were visualized in the metaphase chromosomes of the first cleavage mitosis. From the 2-cell stage on, metaphase chromosomes contained Ag-NORs. Argentophilic proteins were detected in the pronuclei of the 1-cell embryos, i.e. before transcription of the ribosomal genes started. After fertilization these proteins accumulated on the decondensing sperm chromatin and telophase maternal chromosomes, then migrated into the pronuclei to be stored in pronucleoli, and, during mitosis, were transferred into the cytoplasm. In the metaphase chromosomes of the cleaving embryos Ag-NORs adequately reflected the transcriptional activity of the ribosomal genes, whereas in pronuclei of the 1-cell stage embryos argentophilic proteins were not involved in this process, but are likely to play a part in the formation and maturation of pronucleoli, and in the cell cycle regulation.

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PubMed1989

[The ultrastructure and argentophilic properties of the nucleolus organizer and centromeric regions of the chromosomes in early mouse embryogenesis].

The Ag-staining of metaphase chromosomes in one-cell mouse embryos shows that the nucleolus organizer regions (NORs) are Ag-negative, whereas centromeric regions (CRs) are Ag-positive. Starting from 8-16-cell embryos, NORs stained by AgNO3 constantly, CRs remaining argentophobic. On the ultrathin sections of multicell embryos, Ag(+)-NORs differ from the chromosomal arms: they consist of loosely filaments about 6-8 nm in diameter, characterized by a low electron density. On the contrary, at one-cell stage Ag(-)-NORs are not morphologically identified: chromosomal bodies consist of uniform DNP-fibrils about 20 nm in diameter. These data permit to suppose that extended rDNA may form supranucleosomal and nucleosomal DNP-fibrils in the absence of Ag-proteins. The Ag(+)- and Ag(-)-CRs contain 10-20 nm DNP-fibrils mainly, although their density at multicell stages is higher than in one-cell mouse embryos.

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PubMed1988

Cytogenetic study of silver-staining NOR in 8-cell-stage mouse blastomeres fused to 1-cell-stage embryos.

Isolated blastomeres from 8- to 16-cell-stage embryos were fused by standard micromanipulatory means with either unfertilized eggs or fertilized or haploid parthenogenetically activated pronuclear-stage embryos. The hybrid eggs/embryos were incubated overnight in the presence of Colcemid until they had entered the first cleavage division. Air-dried chromosome preparations were then stained with silver nitrate in order to detect active nucleolar organizing regions (NOR). While control unfertilized eggs and 1-cell-stage fertilized and parthenogenetically activated embryos showed no evidence of silver-staining NOR-positive regions, the metaphase plates from 8- to 16-cell embryos showed characteristic NOR-positive regions, while their interphase nuclei also showed a characteristic reticular staining appearance. When hybrids between blastomere nuclei and unfertilized eggs were examined, none of the blastomere nuclei entered mitosis. However, when hybrids between blastomere nuclei and fertilized embryos were examined, in two thirds of the embryos, a single blastomere-derived diploid metaphase plate was present in association with two pronuclear-derived haploid metaphase plates. In most instances, the blastomere-derived chromosomes did not display silver-nitrate-staining NOR. Similar findings were observed when the blastomere-derived chromosomes in hybrids between blastomere nuclei and haploid parthenogenetic embryos were analysed. In the majority of cases, when blastomere nuclei remained in interphase, the characteristic silver-nitrate-staining fine reticular material either was not seen, or the nuclear contents were dispersed into clumps of chromatin-like material. Occasionally, the diploid chromosomes in the hybrids displayed morphological abnormalities. Our findings suggest that the cytoplasm of activated (but not nonactivated) 1-cell embryos is capable of influencing the nucleolar activity of the introduced 8- to 16-cell nuclei, effectively erasing from their chromosomes the memory of at least three previous rounds of rRNA synthesis.

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PubMed1987

[Characteristics of the ultrastructural organization of metaphase chromosomes in mouse embryos in the early cleavage stages].

The structural organization of the mouse metaphase chromosomes in the early embryonic development (I-IV cleavages) was studied using serial ultrathin section. It was shown that in the first cleavage the metaphase chromosomes consist of DNP fibrils 20-25 nm in diameter, which are distributed nonuniformly along the chromosomes. It was suggested that parts of chromosomal arms formed by tightly packing DNP fibrils may correspond to the G-bands revealed by the routine Giemsa staining. In metaphase chromosomes of 8-16-cell embryos DNP fibrils form chromonema--thick threads about 90 nm in diameter. The chromonemata are evenly organized along chromosomal arms. The centromeric heterochromatin always consists of DNP fibrils tightly arranged in a block having no chromonemal level of organization. In all the cells studied chromosomes form structural contacts (associations) by their centromeric heterochromatin regions.

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PubMed1986

[Stimulation and inhibition of the growth of mice carrying the human growth hormone gene].

The human growth hormone gene, containing mouse metallothionein gene promotor, was injected into the male pronuclei of fertilized mouse ova. The progeny of transgenic mice included animals with both accelerated and inhibited growth. Radioimmunochemical analysis has revealed human growth hormone synthesis in both groups of transgenic mice. The molecular weight of the hormone synthesized in liver cells was 25,000 daltons. A possible mechanisms of foreign hormone effect on the growth of transgenic mice is discussed.

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