title: Nuclear-cytoplasmic incompatibility and inefficient development of pig-mouse cytoplasmic hybrid embryos.
abstract: Inter-species somatic cell nuclear transfer (iSCNT) embryos usually fail to develop to the blastocyst stage and beyond due to incomplete reprogramming of donor cell. We evaluated whether using a karyoplast that would require less extensive reprogramming such as an embryonic blastomere or the meiotic spindle from metaphase II oocytes would provide additional insight into the development of iSCNT embryos. Our results showed that karyoplasts of embryonic or oocyte origin are no different from somatic cells; all iSCNT embryos, irrespective of karyoplast origin, were arrested during early development. We hypothesized that nuclear-cytoplasmic incompatibility could be another reason for failure of embryonic development from iSCNT. We used pig-mouse cytoplasmic hybrids as a model to address nuclear-cytoplasmic incompatibility in iSCNT embryos. Fertilized murine zygotes were reconstructed by fusing with porcine cytoplasts of varying cytoplasmic volumes (1/10 (small) and 1/5 (large) total volume of mouse zygote). The presence of pig cytoplasm significantly reduced the development of mouse zygotes to the blastocyst stage compared with control embryos at 120 h post-human chorionic gondotropin (41 vs 6 vs 94%, P<0.05; 1/10, 1/5, control respectively). While mitochondrial DNA copy numbers remained relatively unchanged, expression of several important genes namely Tfam, Polg, Polg2, Mfn2, Slc2a3 (Glut3), Slc2a1 (Glut1), Bcl2, Hspb1, Pou5f1 (Oct4), Nanog, Cdx2, Gata3, Tcfap2c, mt-Cox1 and mt-Cox2 was significantly reduced in cytoplasmic hybrids compared with control embryos. These results demonstrate that the presence of even a small amount of porcine cytoplasm is detrimental to murine embryo development and suggest that a range of factors are likely to contribute to the failure of inter-species nuclear transfer embryos.
authors.affiliation: Animal Development and Biotechnology Group, School of Biosciences, The University of Nottingham, Sutton Bonington, Loughborough, Leicestershire, LE12 5RD, UK.
authors.forename: Dasari
authors.initials: D
authors.lastname: Amarnath
authors.forename: Inchul
authors.initials: I
authors.lastname: Choi
authors.forename: Adel R
authors.initials: AR
authors.lastname: Moawad
authors.forename: Teruhiko
authors.initials: T
authors.lastname: Wakayama
authors.forename: Keith H S
authors.initials: KH
authors.lastname: Campbell
mesh_terms: D000818:Animals; D001755:Blastocyst / Q000502:physiology* / Q000648:ultrastructure; D002417:Cattle; D002459:Cell Fusion / Q000662:veterinary; D002467:Cell Nucleus / Q000378:metabolism*; D019976:Cloning, Organism / Q000379:methods / Q000662:veterinary*; D003593:Cytoplasm / Q000378:metabolism*; D004272:DNA, Mitochondrial / Q000378:metabolism; D046149:Embryo Culture Techniques / Q000662:veterinary; D047108:Embryonic Development; D005260:Female; D020869:Gene Expression Profiling / Q000662:veterinary; D018507:Gene Expression Regulation, Developmental; D008297:Male; D051379:Mice; D009028:Morula / Q000502:physiology / Q000648:ultrastructure; D053652:Nuclear Transfer Techniques / Q000662:veterinary*; D034561:Sheep, Domestic; D013045:Species Specificity; D034421:Sus scrofa; D014158:Transcription, Genetic; D015053:Zygote / Q000502:physiology / Q000648:ultrastructure
chemical_list: D004272:DNA, Mitochondrial