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This Article Full Text Full Text (PDF) All Versions of this Article: 68/4/1249    most recent biolreprod.102.008292v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Ibáñez, E. Articles by Overström, E. W. Search for Related Content PubMed PubMed Citation Articles by Ibáñez, E. Articles by Overström, E. W. Agricola Articles by Ibáñez, E. Articles by Overström, E. W.

Demecolcine-Induced Oocyte Enucleation for Somatic Cell Cloning: Coordination Between Cell-Cycle Egress, Kinetics of Cortical Cytoskeletal Interactions, and Second Polar Body Extrusion¹

^a Department of Biomedical Sciences, Tufts University School of Veterinary Medicine, North Grafton, Massachusetts 01536 ^b Program in Cell, Molecular and Developmental Biology, Department of Anatomy & Cellular Biology, Tufts University School of Medicine, Boston, Massachusetts 02111

Studies were designed to further explore the use of pharmacological agents to produce developmentally competent enucleated mouse oocytes for animal cloning by somatic cell nuclear transfer. Metaphase II oocytes from CF-1 and B6D2F1 strains were activated with ethanol and subsequently exposed to demecolcine at various times postactivation. Chromosome segregation, spindle dynamics, and polar body (PB) extrusion were monitored by fluorescence microscopy using DNA-, microtubule-, and microfilament-selective probes. Exposure to demecolcine did not affect rates of oocyte activation induced by ethanol but did disrupt the coordination of cytokinesis and karyokinesis, suppressing the extent and completion of spindle rotation and second PB extrusion in a strain-dependent manner. Moreover, strain- and treatment-specific variations in the rate of oocyte enucleation were also detected. In particular, CF1 oocytes were more efficiently enucleated relative to B6D2F1 oocytes, and demecolcine treatments initiated early after activation resulted in higher enucleation rates than when treatment was delayed. The observed strain differences are possibly caused by a combination of factors, such as the time course of meiotic cell-cycle progression after ethanol activation, the degree of spindle rotation, and the extent of second PB extrusion. These results suggest that developmentally competent cytoplasts can be produced by timely exposure of activated oocytes to agents that disrupt spindle microtubules. However, the utility of the demecolcine-induced enucleation protocol will require further investigation into factors linking karyokinesis to cytokinesis at the levels of cell-cycle control and oocyte cytoskeletal remodeling following artificial or natural means of egg activation.

¹ Supported in part by USDA-NRI 2001-35205-09966, Charles River Laboratories, and a Fulbright grant sponsored by the Spanish Ministry of Education, Culture & Sports (E.I.).

² Correspondence. FAX: 508 839 7091; eric.overstrom{at}tufts.edu

³ Current address: Departament de Biologia Cel·lular, Fisiologia i Immunologia, Facultat de Ciències, Universitat Autònoma de Barcelona, 08193 Bellaterra, Spain

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				E. Ibanez, D. F Albertini, and E. W Overstrom Effect of genetic background and activating stimulus on the timing of meiotic cell cycle progression in parthenogenetically activated mouse oocytes Reproduction, January 1, 2005; 129(1): 27 - 38. [Abstract] [Full Text] [PDF]