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This Article Full Text Full Text (PDF) All Versions of this Article: 70/3/752    most recent biolreprod.103.024497v1 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 My Folders Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Chang, C.-C. Articles by Tian, X. C. Search for Related Content PubMed PubMed Citation Articles by Chang, C.-C. Articles by Tian, X. C. Agricola Articles by Chang, C.-C. Articles by Tian, X. C.

Nuclear and Microtubule Dynamics of G₂/M Somatic Nuclei During Haploidization in Germinal Vesicle-Stage Mouse Oocytes

Center for Regenerative Biology/Department of Animal Science,² University of Connecticut, Storrs, Connecticut 06269 Reproductive Biology Associates,³ Atlanta, Georgia 30342 Clínica e Centro de Pesquisa em Reprodução Humana Roger Abdelmassih,⁴ São Paulo, Brasil

During the haploidization process, it is expected that diploid chromosomes of somatic cells will be reduced to haploid for the generation of artificial gametes. In the present study, we aimed to use enucleated mouse oocytes at the germinal vesicle-stage (G₂/M) as recipients for somatic cells that are also synchronized to the G₂/M stage for haploidization. The reconstructed oocytes were then induced to undergo meiosis in vitro and observed for their nuclear morphology and microtubule network formation at various expected stages of the meiotic division. Following in vitro maturation, more than half (62/119, 52.1%) of the reconstructed oocytes completed the first round of meiosis-like division, as evidenced by the extrusion of pseudopolar bodies (PBs). However, accelerated PB extrusion, approximately 3–4 h earlier than that by control oocytes occurred. Furthermore, abnormally large pseudo-PBs, as large as four times the normal PB sizes, were observed. During the process of in vitro maturation at both the expected stages of metaphase I (MI) and metaphase II (MII), condensed chromosomes were observed in 38.7% and 55.2% of oocytes, respectively. However, two other types of nuclear configurations were also observed: 1) uneven distribution of chromatin and 2) an interphase-like nucleus, indicating deficiencies in chromosome condensation. Following oocyte activation, more than half (21/33, 63.6%) of the reconstructed oocytes with pseudo-PBs formed separated pseudopronuclei (PN), suggesting formation of functional spindles. The formation of bipolar spindle-like microtubule network at both the expected MI and MII stages during in vitro maturation was confirmed by immunohistochemistry. In summary, this study demonstrated that a high proportion of G₂/M somatic nuclei appear to undergo meiosis-like division, in two successive steps, forming a pseudo-PB and two separate pseudo-PN upon in vitro maturation and activation treatment. Moreover, the enucleated geminal vesicle cytoplast retained its capacity for meiotic division following the introduction of a somatic G₂/M nucleus.

¹ Correspondence: X.C. Tian, Agricultural Biotechnology Laboratory, 1392 Storrs Rd. U 4243, University of Connecticut, Storrs, CT 06269. FAX: 860 486 8809; xtian{at}canr.uconn.edu

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