Adult cloning in cattle: Potential of nuclei from a permanent cell line and from primary cultures

Zakhartchenko, Valeri; Alberio, Ramiro; Stojković, Miodrag; Prelle, Katja; Schernthaner, Wolfgang; Stojković, Petra; Wenigerkind, Hendrik; Wanke, Rüdiger; Duechler, Markus; Steinborn, Ralf; Mueller, Mathias; Brem, Gottfried; Wolf, Eckhard

doi:10.1002/(sici)1098-2795(199911)54:3<264::aid-mrd7>3.0.co;2-y

Cited by 148 publications

(72 citation statements)

References 38 publications

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“…In our results, it should be noted that mean cell numbers per blastocyst in both NT and parthenogenetic embryos was lower than 100 (data not shown), which may indicate delayed and/or abnormal development [3]. Ethanol has been used in cattle to effectively induce oocyte activation [20,21,30] as well as in rabbits [14]. Presicce and Yang [20] demonstrated that a combined ethanol and cycloheximide (a protein synthesis inhibitor) treatment would induce activation of both young (20 hr) and aging (40 hr) IVM bovine oocytes at very high frequencies (90-100%).…”

Section: Discussionmentioning

confidence: 61%

The Effect of Activation Protocols on the Development of Cloned Goat Embryos

Apimeteetumrong

Thuangsanthia²,

Leingcharoen³

et al. 2004

The Journal of Veterinary Medical Science

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ABSTRACT. This study was conducted to compare the developmental competence of somatic nuclear transfer (NT) embryos, after either ionomycin or ethanol activation, in locally bred goats. Donor cells were prepared from the ear skin fibroblasts of a female goat. Cells, at passage 3-8, starved by culturing in 0.5% FCS for 4-8 d, were used for NT. Immature oocytes were obtained from FSH-stimulated goats and matured for 22 hr before enucleation and NT. After fusion, the reconstructed embryos were activated with either ionomycin or ethanol followed by culturing in 6-dimethylaminopurine (6-DMAP) and cytochalasin B (CB), for 3 hr. In experiment I, the fuse d NT embryos (n=63, ionomycin and n=68, ethanol treatments, respectively) were cultured in B 2 with a Vero co-culture system and their developmental competence was evaluated through to Day 9. In experiment II, the NT embryos at the 2-4 cell stage on Day 2 derived from each treatment (ionomycin n=46, and ethanol n=37), were transferred into 10 synchronous recipients. There were no significant differences between the NT embryos derived from the ionomycin and ethanol groups, in fusion (86.3% versus 82.9%), cleavage (90.5% versus 82.4%) and for morula/blastocyst development rates (9.5% versus 5.9%). Sixty percent (3/5) of the recipients from ionomycin became pregnant by midterm (2.5 mts) while only 20% (1/5) from ethanol treatment was pregnant by Day 45. The results demonstrate that activation with either ionomycin or ethanol in combination with 6-DMAP-CB treatment does not affect the development of cloned goat embryos. KEY WORDS: activation, developmental competence, goat, nuclear transfer.J. Vet. Med. Sci. 66(12): 1529-1534, 2004 Since the cloned sheep named "Dolly", was produced by transferring nuclei from adult cells into enucleated oocytes [27], there have been many studies carried out to produce other cloned mammals such as mice, cattle, goats, pigs and rabbits [1,5,19,25,26]. The nuclear transfer (NT) technique has been efficiently used for producing transgenic, cloned farm animal offspring. These animals are capable of producing valuable proteins, which could have a marked impact on the pharmaceutical industry [22] [1,4,6,11,32]. There are a few reports on the development of goat oocytes after parthenogenetic activation by treatment with ionomycin and ethanol, both followed by exposure to 6-diethylaminopurine (6-DMAP) [17,18]. Nevertheless, comparison of the development of NT goat embryos with either ionomycin or ethanol, as an activating agent, has not been reported. The objective of this study was to compare the developmental competence in vitro and in vivo of NT goat embryos, after activation, by using either ionomycin or ethanol, both followed by incubation in 6-DMAP-cytochalasin B (CB). MATERIALS AND METHODSUnless otherwise indicated, all chemicals used in this study were obtained from Sigma-Aldrich Company (St. Louis, MO) and the media from Gibco Invitrogen Corporation (Grand Island, NY).Preparation of donor cells: Donor cells were prepared from an ear skin f...

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Section: Discussionmentioning

confidence: 61%

The Effect of Activation Protocols on the Development of Cloned Goat Embryos

Apimeteetumrong

Thuangsanthia²,

Leingcharoen³

et al. 2004

The Journal of Veterinary Medical Science

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“…The cells were in vitro cultured over several passages and frozen stored before nuclear transfer [2]. However numerous studies provided evidence that somatic cells from a variety of tissues can be used for somatic cell nuclear transfer [12,13].…”

Section: Donor Cellsmentioning

confidence: 99%

Nuclear transfer: a new tool for reproductive biotechnology in cattle

Heyman

2005

Reprod. Nutr. Dev.

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-Recent evolutions of somatic cloning by nuclear transfer are reported, especially in the bovine species where potential applications are underway for biomedicine in association with transgenesis, or for agriculture by improving livestock. The overall efficiency of this biotechnology remains low in terms of viable offspring, but significant progress has been achieved on the different steps of the technique. However, the in vivo development of bovine blastocysts derived from somatic nuclear transfer is characterised by some important features that lead to the "cloning syndrome". Important losses occur during the peri-implantation period and further late foetal loss is observed in association with the Large Offspring Syndrome. About 60-70% of the cloned calves born survive normally to the adult stage and present an apparently normal physiology. Recent data already available on bovine somatic clones of both sexes indicate that they have a zootechnical performance similar to non cloned animals and they are able to reproduce normally without the pathologies associated to cloning thus confirming that the deviations observed in clones are of epigenetic origin and not transmitted to the progeny. biotechnology / reproduction / cattle / somatic cloning / nuclear transfer / efficiency

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“…Exposure to Sr 2+ , which induces multiple calcium transient increases in mouse oocytes, efficiently activated mouse NT embryos that yielded pups [187,188]. In cattle, other treatments that have been used to successfully activate somatic NT embryos include incubation with 7% ethanol [189,190] and cooling to 10 C following oocyte aging [191].…”

Section: Activation Of Nuclear Transfer Embryosmentioning

confidence: 99%

Calcium Release at Fertilization: Artificially Mimicking the Oocyte's Response to Sperm.

Grupen¹,

Nottle²,

Nagashima

2002

Journal of Reproduction and Development

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Abstract. The mechanism of sperm-induced calcium release has been the subject of many studies since the development in the late 1950s of in vitro culture systems that support mammalian fertilization. Despite efforts to elucidate the nature of the signal from the sperm that triggers both the early and late events of oocyte activation, the precise mechanism remains unresolved. Now, with the advent of somatic nuclear transfer technologies, the need to better understand this unique process has been recognised. Nuclear transfer embryos must be induced to commence development artificially because the activating signal from the sperm is absent. The primary activating stimulus is a large increase in the concentration of intracellular-free calcium and numerous physical and chemical treatments have been found to induce calcium changes that initiate the events of oocyte activation. Although live cloned offspring have been produced in a number of species, the overall efficiencies of the nuclear transfer procedures described thus far are unacceptably low and phenotypic anomalies are common. With the aim of improving these efficiencies, researchers are developing artificial activation treatments which induce oocyte responses that mimic those induced by fertilizing sperm. One strategy is to replicate the pattern of calcium change more closely. Another strategy is to couple an activating stimulus with treatments that inhibit maturation (or M-phase) promoting factor (MPF) activity, which regulates meiotic progression in oocytes. This paper reviews what is understood of calcium release at fertilization and describes the treatments that have been used to induce oocyte activation artificially in parthenogenetic and nuclear transfer studies. The relative effectiveness of the strategies employed to mimic the oocyte's response to sperm are discussed. Key words: Oocyte activation, Fertilization, Parthenogenesis, Nuclear transfer (J. Reprod. Dev. 48: 2002) o produce a viable embryo at fertilization, the s p e r m n o t o n l y p r o v i d e s t h e m a l e chromosomal complement, but also the stimulus that initiates development. The mechanism by which the activating signal is transmitted from the sperm to the oocyte is not fully clarified, but it is clear that this signal triggers a large increase in the concentration of intracellular-free calcium. In fertilized sea urchin and Xenopus eggs, the calcium increase takes the form of a propagating wave that originates at the point of sperm attachment and traverses the egg at a velocity of 5 to 10 µm/sec

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Adult cloning in cattle: Potential of nuclei from a permanent cell line and from primary cultures

Cited by 148 publications

References 38 publications

The Effect of Activation Protocols on the Development of Cloned Goat Embryos

The Effect of Activation Protocols on the Development of Cloned Goat Embryos

Nuclear transfer: a new tool for reproductive biotechnology in cattle

Calcium Release at Fertilization: Artificially Mimicking the Oocyte's Response to Sperm.

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