scholarly journals Behaviour of thymocyte nuclei in non-activated and activated mouse oocytes

1984 ◽  
Vol 69 (1) ◽  
pp. 19-34
Author(s):  
R. Czolowska ◽  
J.A. Modlinski ◽  
A.K. Tarkowski
Keyword(s):  
Polyethylene Glycol ◽  
Nuclear Transfer ◽  
Chromosome Condensation ◽  
Premature Chromosome Condensation ◽  
Newborn Mice ◽  
Mouse Oocytes ◽  
Female Pronucleus ◽  
Hybrid Character ◽  
Metaphase Ii Oocytes

Cells originating from the thymus of newborn mice were fused with mouse oocytes using polyethylene glycol. The behaviour of thymocyte nuclei was studied in non-activated metaphase II oocytes, and in oocytes activated in vitro with ethanol. In non-activated oocytes all thymocyte nuclei undergo premature chromosome condensation with individualization of chromosomes; the chromosomes form separate groups in the cytoplasm, or are assembled around the metaphase II spindle, or located on the extra-spindle. In activated oocytes thymocyte nuclei start to develop along a pronucleus-like pathway (decondensation, visualization of nucleoli, swelling) and increase up to 200 times in volume during 24 h culture in vitro, eventually reaching the size of a fully grown pronucleus. Activation/fusion timing seems to be critical for the full remodelling of thymocyte nuclei. Nuclei introduced before (10-30 min) or shortly after (up to 60 min) activation often grow larger than the female pronucleus. Those introduced into oocytes long before activation (greater than 30 min) undergo premature condensation with subsequent reformation of nuclei that are sometimes deficient (as indicated by the presence of micronuclei), or of hybrid character. Nuclei introduced late after activation (greater than 60 min) are mostly doomed to retarded development. The implications of the present observations for nuclear transfer experiments in mammals are discussed.

Progression of mouse oocytes from metaphase I to metaphase II is inhibited by fusion with G2 cells

Zygote ◽  
2000 ◽  
Vol 8 (2) ◽  
pp. 145-151 ◽  
Author(s):  
Joanna Grabarek ◽  
Magdalena Zernicka-Goetz
Keyword(s):  
Chromosome Condensation ◽  
Factor Activity ◽  
Premature Chromosome Condensation ◽  
Mouse Oocytes ◽  
M Phase ◽  
G2 Cells ◽  
Cytostatic Factor ◽  
Metaphase Ii Oocytes ◽  
Metaphase I

We show that in contrast to metaphase II oocytes, metaphase I oocytes cannot be activated by fusion with the zygote. Fusion of metaphase I oocytes with G2 zygotes was followed by premature chromosome condensation, with 60% of the hybrids becoming arrested at metaphase I, the remainder progressing and arresting at metaphase II. Hybrids of metaphase I oocytes and M-phase zygotes underwent accelerated maturation, but all arrested at metaphase II. In both cases the arrest could be overcome by treatment with the parthenogenetic activators ethanol and cycloheximide. We discuss these findings in relation to the possibility that the metaphase I oocyte contains cytostatic factor activity that is activated by its zygotic partner. Alternatively, the G2 zygote may provide an inhibitor of anaphase, normally never present in the metaphase I oocyte and which is absent from the M-phase zygote.

In vitro fertilisation of mouse oocytes reconstructed by transfer of metaphase II chromosomes results in live births

Zygote ◽  
2001 ◽  
Vol 9 (1) ◽  
pp. 9-14 ◽  
Author(s):  
Min-Kang Wang ◽  
Da-Yuan Chen ◽  
Ji-Long Lui ◽  
Guang-Peng Li ◽  
Qing-Yuan Sun
Keyword(s):  
Nuclear Transfer ◽  
Normal Structure ◽  
Human Reproduction ◽  
In Vitro Fertilisation ◽  
Kunming Mouse ◽  
Mouse Oocytes ◽  
Assisted Human Reproduction ◽  
And Function ◽  
Apparatus Transfer

The interaction between nucleus and cytoplasm can be explored through nuclear transfer. We describe here another tool to investigate this interaction: MII meiotic apparatus transfer (MAT) between mouse oocytes. In this study, the MII oocyte meiotic apparatus or spindle from C57BL/6 mice, a black strain, was transferred into an enucleated metaphase oocyte from Kunming mouse, a white strain. The results showed that the enucleation rate by treating oocytes with 3% sucrose was 100%, but the electrofusion efficiency was very low, with only 17.6% of reconstructed karyoplast-recipient cytoplasm pairs fused. When the fused oocytes were exposed to spermatozoa from C57BL/6 mice, 9 of 11 (82%) were fertilised. Eight reconstructed embryos at 1- to 4-cell stages were transferred into the oviducts of two synchronously pregnant Kunming strain fosters and one delivered two normal C57BL/6 offspring. This study indicates that MII meiotic apparatus or spindle sustains normal structure and function after micromanipulation and electrofusion. MAT provides a model for further research on the application of this technique to assisted human reproduction.

Nuclear envelope removal/maintenance determines the structural and functional remodelling of embryonic red blood cell nuclei in activated mouse oocytes

Zygote ◽  
1998 ◽  
Vol 6 (1) ◽  
pp. 65-73 ◽  
Author(s):  
Daniel Szöllösi ◽  
Renata Czołowska ◽  
Ewa Borsuk ◽  
Maria S. Szöllösi ◽  
Pascale Debey
Keyword(s):  
Nuclear Envelope ◽  
Transcriptional Activity ◽  
Chromatin Condensation ◽  
Oocyte Activation ◽  
Cell Nuclei ◽  
Nuclear Envelope Breakdown ◽  
Mouse Oocytes ◽  
Female Pronucleus ◽  
Mouse Fetuses

SummaryNuclei of embryonic red blood cells (e-RBC) from 12-day mouse fetuses are arrested in Go phase of the cell cycle and have low transcriptional activity. These nuclei were transferred with help of polyethylene glycol (PEG)-mediated fusion to parthenogenetically activated mouse oocytes and heterokaryons were analysed for nuclear structure and transcriptional activity. If fusion proceeded 25–45 min after oocyte activation, e-RBC nuclei were induced to nuclear envelope breakdown and partial chromatin condensation, followed by formation of nuclei structurally identical with pronuclei. These ‘pronuclei’, similar to egg (female) pronuclei, remained transcriptionally silent over several hours of in vitro culture. If fusion was performed 1 h or later (up to 7 h) after activation, the nuclear envelope of e-RBC nuclei remained intact and nuclear remodelling was less spectacular (slight chromatin decondensation, formation of nucleolus precursor bodies). These nuclei, however, reinforced polymerase-II-dependent transcription within a few hours of in vitro culture. Our present experiments, together with our previous work, demonstrate that nuclear envelope breakdown/maintenance are critical events for nuclear remodelling in activated mouse oocytes and that somatic dormant nuclei can be stimulated to renew transcription at a time when the female pronucleus remains transcriptionally silent.

Behaviour of mouse primary spermatocyte nuclei after fusion to enucleated metaphase II oocytes

Zygote ◽  
2000 ◽  
Vol 8 (4) ◽  
pp. 295-302 ◽  
Author(s):  
Michal Kubelka ◽  
Robert M. Moor
Keyword(s):  
Cell Division ◽  
Light Microscopy ◽  
Dna Synthesis ◽  
Chemical Treatment ◽  
Morphological Changes ◽  
Primary Spermatocyte ◽  
Chromosome Condensation ◽  
Premature Chromosome Condensation ◽  
Similar Morphology ◽  
Metaphase Ii Oocytes

Primary spermatocytes originating from prepubertal mouse testes were electrofused to metaphase II (MII)-stage oocytes, enucleated either by the conventional micromanipulation method or by chemical treatment with etoposide and cycloheximide. These experiments were followed by assessment of morphological changes in transferred nuclei using light microscopy, by chromosomal analyses and by screening of hybrids for the presence or absence of DNA synthesis using anti-bromodeoxyuridine antibody and immunofluorescence staining of the hybrids. The results show differences between the two types of ooplasts in susceptibility to activation stimuli. However, when activated, both types of ooplasts gave rise to hybrids of similar morphology. From 35.3% to 63% of activated hybrids originating from chemically or microsurgically enucleated oocytes, respectively, contained one large pronucleus in cytoplasm, 62% or 31.6% hybrids from those two groups, respectively, possessed two smaller pronuclei and a few contained three or four pronuclei. No DNA synthesis was detected in any hybrid containing one or more pronuclei. The chromosome spreads of hybrids with premature chromosome condensation (PCC) morphology (before activation) show that most of the hybrids had a diploid (2n) number of chromosomes. The nature and regularity of the cell division cycle in the hybrids are discussed.

Chromosome condensation activity in ovulated metaphase II mouse oocytes assayed by fusion with interphase blastomeres

1986 ◽  
Vol 84 (1) ◽  
pp. 129-138
Author(s):  
R. Czolowska ◽  
M. Waksmundzka ◽  
J.Z. Kubiak ◽  
A.K. Tarkowski
Keyword(s):  
Interphase Nucleus ◽  
Volume Ratio ◽  
Chromosome Condensation ◽  
Chorionic Gonadotrophin ◽  
Human Chorionic Gonadotrophin ◽  
Experimental Protocol ◽  
Premature Chromosome Condensation ◽  
Mouse Oocytes ◽  
Condensation Activity ◽  
Fusion Products

Fusion of large and small karyoplasts produced from metaphase II mouse oocytes with interphase blastomeres from 2-cell and 8-cell embryos (volume ratio of partners, 1:1) results in premature chromosome condensation (PCC) of the interphase nucleus in the majority of the fusion products (hybrids). Fused under the same experimental protocol, oocyte-derived cytoplasts also induce PCC of the blastomere nucleus in the fusion products (cybrids) provided they originate from recently ovulated oocytes (141/2-15 h after injection of human chorionic gonadotrophin (HCG)). In cytoplasts derived from older oocytes (16–20 h post-HCG) chromosome condensation activity gradually decreases with time as can be inferred from the increasing proportion of cybrids retaining interphase blastomere nuclei. However, even the oldest cytoplasts (19–20 h post-HCG) can induce PCC if the cytoplast volume significantly exceeds the volume of the interphase partner (7:1). We postulate that the condensation activity is predominantly bound to the nuclear apparatus (most probably to the chromosomes), and that in the cytoplasm of metaphase II mouse oocyte it decreases with post-ovulatory age.

Oocyte age and nuclear donor cell type affect the technical efficiency of somatic cloning in rabbits

Zygote ◽  
2003 ◽  
Vol 11 (2) ◽  
pp. 151-158 ◽  
Author(s):  
Rita P. Cervera ◽  
Fernando García-Ximénez
Keyword(s):  
Technical Efficiency ◽  
Nuclear Transfer ◽  
Donor Cell ◽  
Cumulus Cells ◽  
Cell Types ◽  
Cell Type ◽  
Fetal Fibroblasts ◽  
Somatic Nuclear Transfer ◽  
Metaphase Ii Oocytes

The present study in rabbits compared, in the first experiment, the effect of two commonly used oocyte ages, 13 h and 17 h after ovulation induction treatment, on the technical efficiency of somatic nuclear transfer steps, using fresh cumulus cells as nuclear donors. Recently ovulated metaphase II oocytes (13 h) showed higher fusion (13 h: 83% vs 17 h: 67%, p < 0.05) and in vitro development rates than in vivo slightly aged metaphase II oocytes (morula, 13 h: 74% vs 17 h: 25%, p < 0.05; blastocyst, 13 h: 16% vs 17 h: 8%; p < 0.05). In contrast, activation rate was higher in the 17 h group (13 h: 45% vs 17 h: 67%; p < 0.05). In a second experiment, using recently ovulated oocytes (13 h) as recipients, two donor cell types (from primary cultures of either cumulus cells or fetal fibroblasts) were tested to evaluate their effects on the efficiencies of the different technical steps of somatic nuclear transfer procedure. A better fusion rate was obtained when fetal fibroblasts were used as nuclear donors (cumulus cells: 45% vs fetal fibroblasts: 67%, p < 0.05). No statistically significant differences were detected in cleavage rate regardless of the cell type used (cumulus cells: 44% vs fetal fibroblasts: 60%, p > 0.05). However, in vitro development to morula (cumulus cells: 41% vs fetal fibroblasts: 14%, p < 0.05) and to blastocyst stage (cumulus cells: 27% vs fetal fibroblasts: 3%, p < 0.05) were different between cell types.

Meiotic maturation of mouse oocytes in vitro: association of newly synthesized proteins with condensing chromosomes

1976 ◽  
Vol 20 (3) ◽  
pp. 549-568 ◽  
Author(s):  
P.M. Wassarman ◽  
G.E. Letourneau
Keyword(s):  
Protein Synthesis ◽  
Mammalian Species ◽  
Germinal Vesicle ◽  
Chromosome Condensation ◽  
Intracellular Distribution ◽  
Meiotic Maturation ◽  
Meiotic Progression ◽  
Mouse Oocytes

The nature, intracellular distribution, and role of proteins synthesized during meiotic maturation of mouse oocytes in vitro have been examined. Proteins synthesized during the initial stages of maturation are concentrated within the nucleus (germinal vesicle) and become intimately associated with the condensing chromosomes. Inhibition of protein synthesis during this period does not prevent germinal vesicle dissolution or chromosome condensation, but meiotic progression is blocked reversibly at the circular bivalent stage. A protein is synthesized during meiotic maturation of the mouse oocyte which exhibits several of the characteristics of the very lysine-rich histone, FI; this and other histones are phosphorylated during the initial stages of maturation. These results are discussed in relation to studies of meiotic maturation of oocytes from non-mammalian species and chromosome condensation in both oocytes and mitotic cells.

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