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PubMed2004

Activation and early parthenogenesis of bovine oocytes treated with ethanol and strontium.

Efficient artificial activation is indispensable for the success of cloning programs. Strontium has been shown to effectively activate mouse oocytes for nuclear transfer procedures, however, there is limited information on its use for bovine oocytes. The present study had as objectives: (1). to assess the ability of strontium to induce activation and parthenogenetic development in bovine oocytes of different maturational ages in comparison with ethanol; and (2). to verify whether the combination of both treatments improves activation and parthenogenetic development rates. Bovine oocytes were in vitro matured for 24, 26, 28, and 30 h, and treated with ethanol (E, 7% for 5 min) or strontium chloride (S, 10mM SrCl(2) for 5h) alone or in combination: ethanol+strontium (ES) and strontium+ethanol (SE). Activated oocytes were cultured in vitro in synthetic oviductal fluid (SOF) medium and assessed for pronuclear formation (15-16 h), cleavage (46-48 h) and development to the blastocyst stage (D7). Treatment with ethanol and strontium promoted similar results regarding pronuclear formation (E, 20-66.7%; S, 26.7-53.3%; P>0.05) and cleavage (E, 12.8-40.6%; S, 16.1-41.9%; P>0.05), regardless of oocyte age. The actions of both strontium and ethanol were influenced by oocyte age: ethanol induced greater activation rates after 28 and 30 h of maturation (48.4 and 66.7% versus 20.0 and 23.3% for 24 and 26 h, respectively; P<0.05) and strontium after 30 h (53.3%) was superior to 24 and 26 h (26.7% for both). Blastocyst development rates were minimal in all treatments (0.0-6.3%; P>0.05), however, when the mean (+/-S.D.) cell number in blastocysts at the same maturational period was compared, strontium treatment was superior to ethanol for activation rates (82+/-5.7 and 89.5+/-7.8 versus 54 and 61, at 28 and 30 h, respectively). Improved results were obtained by combined treatments. The combination of ethanol and strontium resulted in similar pronuclear formation (ES, 36.7-83.9%; SE, 53.1-90.3%) and cleavage rates (ES, 31.3-81.3%; SE, 65.6-80.7%). Regarding embryo development, there was no difference (P>0.05) between treatments, and blastocysts were only obtained in treatment SE at 24 and 26 h (6.5% for both). It is concluded that, SrCl(2) induces activation and parthenogenetic development in bovine oocytes.

Animals↗
PubMedDate not supplied

[From human andro- and parthenogenesis (hydatidiform moles and benign ovarian teratomas) to cancer].

Genomic imprinting is a process that appeared in mammals. This phenomenon blocks the normal development of parthenogenic and androgenic conceptuses, that is to say benign ovarian teratomas and hydatidiform moles respectively. Pathological modifications of these conceptuses depend on whether the chromosomes come from the mother or father. These pathologies are associated with an accidental anomaly during gametogenesis and/or fertilizing. These reproductive anomalies are sporadic and some familial cases may exist suggesting a genetic control of such diseases. The human andro- and parthenogenetic conceptuses, but more frequently the moles, may be invasive (choriocarcinoma). An imbalance of the imprinting genes may initiate the deregulation of other genes, including oncogenes and anti-oncogenes, which can explain the cancerous modification. Immunological and environmental factors must be also considered (presence of the only paternal chromosomes in the choriocarcinoma). Numerous works on this subject are published and some recent important discoveries underline the roles of genes HOX, Tim P3, E-cad and p-16, and the recurrent chromosome anomalies 7q21+and 8p21- in the mole to choriocarcinoma processing. Although these phenomena are complex and heterogeneous, the andro- and parthenogenote conceptuses are particularly interesting models with which to understand developmental disorders and cancerous progression.

Embryonic Development↗
PubMed2003

Fertilized eggs obtained from transplantation of frozen ovaries and parthenogenesis in combination with artificial insemination of frozen semen of the silkworm, Bombyx mori.

A reliable method is reported for the long-term preservation of ovaries and spermatozoa of the silkworm (Bombyx mori). Three studies are presented. In the first, ovaries were removed from larvae at either 3rd, 4th, or 5th instar, cryopreserved, and stored in liquid nitrogen. Thawed ovaries were transplanted to surgically castrated female larvae at the same or a different developmental stage. The highest percentage of recipient females producing eggs resulted into either 3rd or 4th instar larvae (respectively, 22.1 and 8.7%). Similarly, the highest levels of other measurements of successful cryopreservation and transplanted ovary, and number of eggs laid, occurred with the same combination of donor and recipient developmental stages. Other combinations of ovary/recipient developmental stages yielded lower results. In the second experiment, semen was collected from male moths, cryopreserved, and then thawed semen was diluted with trypsin solution and artificially inseminated into females obtained from the best conditions of first experiment. A small percentage of inseminated moths laid eggs (8-10.3%) compared to that of controls (100%). In addition, the fertility of eggs from experimental moths was lower than that of control females (respectively, 40.3-88% and 97.5%). In the third experiment, eggs were surgically removed from ovarian tubules of moth following transplantation of thawed ovaries and subjected to parthenogenetic activation and artificial hatching. As expected, all resulting moths were female and, following natural mating or artificial insemination with thawed semen, yielded normal offspring at high rates.

Animals↗
PubMed2003

Drosophila parthenogenesis: a model for de novo centrosome assembly.

The Drosophila egg contains all the components required to properly execute the early mitotic divisions but is unable to assemble a functional centrosome without a sperm-provided basal body. We show that 65% of unfertilized eggs obtained from a laboratory strain of Drosophila mercatorum can spontaneously assemble a number of cytoplasmic asters after activation, most of them duplicating in a cell cycle-dependent manner. Such asters are formed by a polarized array of microtubules that have their Asp-associated minus-ends converging at a main focus, where centrioles and typical centrosomal antigens are found. Aster assembly is spatially restricted to the anterior region of the oocyte. When fertilized, the parthenogenetic egg forms the poles of the gonomeric spindle by using the sperm-provided basal body, despite the presence within the same cytoplasm of maternal centrosomes. Thirty-five percent of parthenogenetic eggs and all unfertilized and fertilized eggs from the sibling bisexually reproducing D. mercatorum strain do not contain cytoplasmic asters. Thus, the Drosophila eggs have the potential for de novo formation of functional centrosomes independent of preexisting centrioles, but some control mechanisms preventing their spontaneous assembly must exist. We speculate that the release of the block preventing centrosome self-assembly could be a landmark for ensuring parthenogenetic reproduction.

Animals↗
PubMed2002

Parthenogenesis of rabbit oocytes activated by different stimuli.

Oocyte activation is one of the essential elements determining the success of nuclear transfer and the subsequent development of cloned embryos both in vitro and in vivo. Experiments were conducted to optimize the protocol for oocyte activation in a regular nuclear transfer study. In vivo derived oocytes were collected at 14-15 h from New Zealand white rabbits after ovulation treatment and were activated +18 h post-ovulation treatment. Single activation agents including calcium ionophore (A23187, 5 microM, 5 min), ethanol (Eth, 7%, 7 min), and thimerosal (200 microM, 10 min) were tested. Cleavage rates were highest in the ethanol-treated group (37%) compared to other treatments (19-25%). Very low blastocyst rates (2-3%) resulted which were not significantly different among treatments (P>0.05). Combined single agent treatment (calcium stimulators) with protein kinase inhibitor, 6-DMAP were used to achieve a full oocyte activation. Both pronuclear and blastocyst formation rates were significantly higher (P<0.05) in the Eth+6-DMAP treatment group (38 and 27%) than in the other groups (16-21 and 7-9%, respectively, P<0.05). Low (0.2mM) and high (2.5mM) concentrations of 6-DMAP treatments with different treatment lengths (1.5 and 3.5h) in the combined groups were also compared. Blastocyst formation and cleavage rates were greater in the high concentration with less treatment time groups (36% versus 4-20%, P<0.05). In conclusion, single activation agents, either Ca2+ stimulators or protein kinase inhibitors, could not fully activate mature rabbit oocytes. The best activation procedure obtained in this study was the Eth+6-DMAP combined treatment, which may be incorporated into regular nuclear transfer or cloning protocols.

Adenine↗