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Nuclear-Cytoplasmic Interactions Affect In Utero Developmental Capacity, Phenotype, and Cellular Metabolism of Bovine Nuclear Transfer Fetuses¹

Department of Molecular Animal Breeding and Biotechnology,⁴ Gene Center of the Ludwig-Maximilian University, D-81377 Munich, Germany Department of Animal Breeding and Genetics and Central Biotechnical Unit Strahlenzentrum,⁵ Justus-Liebig-University, D-35392 Giessen, Germany Bavarian Research Station for Animal Breeding,⁶ D-85586 Grub, Germany Bavarian Research Center for Biology of Reproduction (BFZF),⁷ D-85764 Oberschleissheim, Germany Agrobiogen GmbH,⁸ D-86567 Hilgertshausen, Germany

We generated a clone of bovine somatic cell nuclear transfer embryos using oocyte pools from defined maternal sources to study nuclear-cytoplasmic interactions. Nucleocytoplasmic hybrids were reconstructed with Bos taurus (Brown Swiss) granulosa cells and oocytes that contained B. taurus A (Simmental), B. taurus B (Simmental), or Bos indicus (Dwarf Zebu) cytoplasm. Another set of embryos was reconstructed with randomly selected Brown Swiss (B. taurus R) oocytes. Embryo transfer resulted in nine (12.5%), nine (13.8%), three (50%), and 11 (16.7%) Day 80 fetuses, of which eight (11.1%), three (4.6%), three (50%), and 10 (15.2%) were viable, respectively. The proportion of viable fetuses was affected by cytoplasm (likelihood ratio test, P < 0.02) and was higher for embryos with B. indicus cytoplasm than for the B. taurus A (P < 0.05) and B (P < 0.01) groups. Furthermore, the proportion of surviving Day 80 fetuses was reduced for B. taurus B as compared with B. taurus A and B. taurus R cytoplasm (P < 0.05 and P < 0.02). Body weight of nucleocytoplasmic hybrid fetuses was not significantly different from Brown Swiss control fetuses produced by artificial insemination (AI), but fetuses reconstructed with random cytoplasts of the same breed as the nuclear donor exhibited overgrowth (P < 0.01) and a higher coefficient of variation in weight. Furthermore, body weight, crown rump length, thorax circumference (P < 0.05), and femur length (P < 0.01) of fetuses with B. taurus A cytoplasm differed from fetuses with B. taurus R cytoplasms. Fetal skin, heart, and liver cells with B. indicus cytoplasm showed a greater increase in number per time period (P < 0.001) and oxygen consumption rate per cell (skin and liver, P < 0.001; heart, P < 0.08) in comparison with their counterparts with B. taurus A cytoplasm. These data point to complex oocyte cytoplasm-dependent epigenetic modifications and/or nuclear DNA-mitochondrial DNA interactions with relevance to nuclear transfer and other reproductive technologies such as ooplasmic transfer in human assisted reproduction.

¹ Supported by grants from the Deutsche Forschungsgemeinschaft (HI 503/3-1, WO 685/3-1).

² Correspondence: Stefan Hiendleder, Department of Molecular Animal Breeding and Biotechnology, Ludwig-Maximilian University Munich, Hackerstrasse 27, D-85764 Oberschleissheim, Germany. FAX: 49 89 315 2799; s.hiendleder{at}gen.vetmed.uni-muenchen.de

³ Permanent address: Department of Animal Biology, University of Sassari, 07100 Sassari, Italy

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