Parthenogenetic development of bovine oocytes treated with ethanol and cytochalasin b after in vitro maturation

Fukui, Yutaka; Sawai, Ken; Furudate, Makoto; Sato, Noriyuki; Iwazumi, Yasuaki; Ohsaki, Kazue

doi:10.1002/mrd.1080330318

Cited by 95 publications

(70 citation statements)

References 20 publications

(25 reference statements)

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“…This resulted in diploid zygotes with two pronuclei (14,15,17,35,37). The present results show that CB did not affect chromosomal movement or nuclear division but did inhibit spindle rotation and, thus, cytokinesis.…”

Section: Discussionmentioning

confidence: 50%

“…The histone kinase inhibitor 6-dimethylaminopurine has been reported to accelerate and enhance the formation of pronuclei in non-aged MII oocytes from mice and cattle (16). Also, cytochalasin B (CB), an inhibitor of microfilament polymerization, prevents the release of the second polar body (Pb2) after oocyte activation, which would result in diploid development (17), and may also help prevent embryo fragmentation (18,19 (22)(23)(24). Therefore, it is important to investigate [Ca 2+ ] i oscillations of oocytes treated with CM when judging the efficiency of CM for oocyte activation and embryo development.…”

Section: Introductionmentioning

confidence: 99%

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Effects of the conditioned medium of mesenchymal stem cells on mouse oocyte activation and development

Feng

Zhou

Ling

et al. 2009

Braz J Med Biol Res

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Mesenchymal stem cells (MSCs) have been reported to secrete a variety of cytokines and growth factors acting as trophic suppliers, but little is known regarding the effects of conditioned medium (CM) of MSCs isolated from femurs and tibias of mouse on the artificial activation of mouse oocytes and on the developmental competence of the parthenotes. In the current study, we investigated the effect of CM on the events of mouse oocyte activation, namely oscillations of cytosolic calcium concentration ([Ca 2+ ] i ), meiosis resumption, pronucleus formation, and parthenogenetic development. The surface markers of MSCs were identified with a fluorescence-activated cell sorter. The dynamic changes of the spindle and formation of pronuclei were examined by laser-scanning confocal microscopy. Exposure of cumulus-oocyte complexes to CM for 40 min was optimal for inducing oocyte parthenogenetic activation and evoking [Ca 2+ ] i oscillations similar to those evoked by sperm (95 vs 100%; P > 0.05). Parthenogenetically activated oocytes immediately treated with 7.5 μg/mL cytochalasin B (CB), which inhibited spindle rotation and second polar body extrusion, were mostly diploid (93 vs 6%, P < 0.01) while CB-untreated oocytes were mostly haploid (5 vs 83%, P < 0.01). Consequently, the blastocyst rate was higher in the CB-treated than in the CB-untreated oocytes. There was no significant difference in developmental rate between oocytes activated with CM and 7% ethanol (62 vs 62%, P > 0.05), but the developmental competence of the fertilized oocytes was superior to that of the parthenotes (88 vs 62%, P < 0.05). The present results demonstrate that CM can effectively activate mouse oocytes, as judged by the generation of [Ca 2+ ] i oscillations, completion of meiosis and parthenogenetic development.

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Section: Discussionmentioning

confidence: 50%

Section: Introductionmentioning

confidence: 99%

Effects of the conditioned medium of mesenchymal stem cells on mouse oocyte activation and development

Feng

Zhou

Ling

et al. 2009

Braz J Med Biol Res

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“…They resume the mitotic cell cycle due to subsequent exposure to protein synthesis inhibitors [7] or broad-spectrum protein kinase inhibitors [30]. Enhancement of the developmental competence of bovine oocytes has been reported by the use of cytochalasin B (CCB), which may prevent extrusion of the second PB, resulting in diploid development [9]. The present study was designed to evaluate the effect of Ion, CHX and the combination of CHX with CCB on the developmental rates and ploidy of bovine parthenote embryos.…”

Section: Discussionmentioning

confidence: 99%

Parthenogenetic Development and Ploidy following Various Chemical Activation Regiments of Bovine Oocytes

Ock

Rho

2008

The Journal of Veterinary Medical Science

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ABSTRACT. Bovine oocytes treated using various combinations of 5 µM ionomycin (Ion), 10 µg/ml cycloheximide (CHX) and 5 µg/ml cytochalsin B (CCB) were evaluated to determine developmental rates and ploidy status. The groups were comprised of metaphase II oocytes exposed to Ion for 5 min (Group 1); Ion treatment followed by CHX for 5 hr (Group 2); Ion treatment followed by CHX/CCB for 1 hr and CHX for 4 hr (Group 3); Ion treatment followed by CHX/CCB for 3 hr and CHX for 2 hr (Group 4); and Ion treatment followed by CHX/CCB for 5 hr (Group 5). Group 5 exhibited significantly (P<0.05) lower rates of second polar body (PB) extrusion and higher rates of cleavage and blastocyst development. At 8 hr after Ion treatment, the eggs in groups 2, 3 and 5 were divided into 2 subgroups based on the presence or absence of the second PB and were assessed for cleavage rate and ploidy at the two-cell stage. The cleavage rates did not differ among the activation treatments or between the presence and absence of the second PB in all groups. The diploid rate was significantly (P<0.05) higher in group 5 than in groups 2 and 3. However, the diploid rate in blastocyst-stage parthenotes did not differ among groups 2, 3 and 5. Consequently, oocyte activation by CHX/CCB for 5 hr after Ion treatment could enhance diploid parthenogenetic development in bovine. KEY WORDS: bovine parthenote, chemical activation, ploidy, polar body.J. Vet. Med. Sci. 70(11): 1165-1172, 2008 In bovine, oocyte activation is a major factor for successful production of reconstructed embryos by somatic cell nuclear transfer (SCNT) [34], parthenogenesis and intracytoplasmic sperm injection [23]. During normal fertilization in mammalian species, sperm penetration into the oocyte triggers intracellular Ca 2+ oscillation followed by inactivation of the maturation promoting factor (MPF) composed of the catalytic subunit of cdc2p34 protein kinase and the regulatory subunit cyclin B1/B2 for about 2 hr [37] [29,30], that prevent MPF re-accumulation by reducing cdc2 kinase activity [2] have also been applied. DMAP treatment of metaphase II oocytes induces diploid activation by preventing chromosomal separation and extrusion of the second polar body (PB) [19]. However, CHX does not prevent chromosomal segregation and second PB extrusion, resulting in haploid development. When cytochalasin D (CCD) was added in the medium, chromosomal segregation manifested, but the cytokinesis that induces second PB extrusion was prohibited in CHX-treated oocytes, leading to diploid development with two pronuclei in activated oocytes [4,19]. The combined treatment of ionomycin with DMAP for oocyte activation could induce an abnormal pattern of karyokinesis during the first cell cycle [8], and thus, CHX has widely been used as an alternative approach [19]. Moreover, in cloning, oocyte activation using CHX produces healthy live offspring in cattle [12,17], but the live offspring rate is still low [5].Campbell et al. [3] proposed the hypothesis that when donor cells at G1 stage are reconstructed...

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“…Essential alterations in parthenotes affect not only the trophoblast (something precociously evident at the blastocyst stage), but also parts of the embryo proper, such as muscle, liver, and pancreas (Fundele et al 1989(Fundele et al , 1990. In the cow, the maximum development in uterus reached by a parthenogenetic embryo has been reported to be of 48 days (Fukui et al 1992). The arrest in development of parthenotes is thought to be due to alterations in their genomic imprinting, which in turn seems to be an evolutive result of conflicts between the parental genomes during growth (Brevini & Gandolfi 2008).…”

Section: Introductionmentioning

confidence: 99%

Biological differences between in vitro produced bovine embryos and parthenotes

Gómez¹,

Gutiérrez‐Adán²,

Díez³

et al. 2009

Reproduction

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Parthenotes may represent an alternate ethical source of stem cells, once biological differences between parthenotes and embryos can be understood. In this study, we analyzed development, trophectoderm (TE) differentiation, apoptosis/necrosis, and ploidy in parthenotes and in vitro produced bovine embryos. Subsequently, using real-time PCR, we analyzed the expression of genes expected to underlie the observed differences at the blastocyst stage. In vitro matured oocytes were either fertilized or activated with ionomycin C6-DMAP and cultured in simple medium. Parthenotes showed enhanced blastocyst development and diploidy and reduced TE cell counts. Apoptotic and necrotic indexes did not vary, but parthenotes evidenced a higher relative proportion of apoptotic cells between inner cell mass and TE. The pluripotence-related POU5F1 and the methylation DNMT3A genes were downregulated in parthenotes. Among pregnancy recognition genes, TP-1 was upregulated in parthenotes, while PGRMC1 and PLAC8 did not change. Expression of p66 shc and BAX/BCL2 ratio were higher, and p53 lower, in parthenotes. Among metabolism genes, SLC2A1 was downregulated, while AKR1B1, PTGS2, H6PD, and TXN were upregulated in parthenotes, and SLC2A5 did not differ. Among genes involved in compaction/blastulation, GJA1 was downregulated in parthenotes, but no differences were detected within ATP1A1 and CDH1. Within parthenotes, the expression levels of SLC2A1, TP-1, and H6PD, and possibly AKR1B1, resemble patterns described in female embryos. The pro-apoptotic profile is more pronounced in parthenotes than in embryos, which may differ in their way to channel apoptotic stimuli, through p66 shc and p53respectively, and in their mechanisms to control pluripotency and de novo methylation.Reproduction (2009) 137 285-295

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Parthenogenetic development of bovine oocytes treated with ethanol and cytochalasin b after in vitro maturation

Cited by 95 publications

References 20 publications

Effects of the conditioned medium of mesenchymal stem cells on mouse oocyte activation and development

Effects of the conditioned medium of mesenchymal stem cells on mouse oocyte activation and development

Parthenogenetic Development and Ploidy following Various Chemical Activation Regiments of Bovine Oocytes

Biological differences between in vitro produced bovine embryos and parthenotes

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