The aim of this study was to determine whether the number of passages affected the developmental pluripotency of embryonic stem (ES) cells as measured by the attainment of adult fertile mice derived from embryonic stem (ES) cell/tetraploid embryo complementation. Thirty-six newborns were produced by the aggregation of tetraploid embryos and hybrid ES cells after various numbers of passages. These newborns were entirely derived from ES cells as judged by microsatellite DNA, coat-color phenotype, and germline transmission. Although 15 survived to adulthood, 17 died of respiratory failure, and four were eaten by their foster mother. From the 15 mice that reached adulthood and that could reproduce, none arose from ES cells at passage level 15 or more. All 15 arose from cells at passages 3–11. Our results demonstrate that the number of passages affects the developmental pluripotency of ES cells.
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Boyd S (2004) Causes and treatment of neonatal respiratory distress syndrome. Nurs Times 100:40–44
Brook FA, Gardner RL (1997) The origin and efficient derivation of embryonic stem cells in the mouse. Proc Natl Acad Sci USA 94:5709–5712
Dean W, Bowden L, Aitchison A, Klose J, Moore T, Meneses JJ, Reik W, Feil R (1998) Altered imprinted gene methylation and expression in completely ES cell-derived mouse fetuses: association with aberrant phenotypes. Development 125:2273–2282
Eakin GS, Behringer RR (2003) Tetraploid development in the mouse. Dev Dyn 228:751–766
Eakin GS, Hadjantonakis AK, Papaioannou VE, Behringer RR (2005) Developmental potential and behavior of tetraploid cells in the mouse embryo. Dev Biol 288:150–159
Eggan K, Jaenisch R (2003) Differentiation of F1 embryonic stem cells into viable male and female mice by tetraploid embryo complementation. Methods Enzymol 365:25–39
Eggan K, Akutsu H, Loring J, Jackson-Grusby L, Klemm M, Rideout WM, Yanagimachi R, Jaenisch R (2001) Hybrid vigor, fetal overgrowth, and viability of mice derived by nuclear cloning and tetraploid embryo complementation. Proc Natl Acad Sci USA 98:6209–6214
Eggan K, Rode A, Jentsch I, Samuel C, Hennek T, Tintrup H, Zevnik B, Erwin J, Loring J, Jackson-Grusby L, Speicher MR, Kuehn R, Jaenisch R (2002) Male and female mice derived from the same embryonic stem cell clone by tetraploid embryo complementation. Nat Biotechnol 20:455–459
Eggan K, Baldwin K, Tackett M, Osborne J, Gogos J, Chess A, Axel R, Jaenisch R (2004) Mice cloned from olfactory sensory neurons. Nature 428:44–49
Hochedlinger K, Jaenisch R (2002) Monoclonal mice generated by nuclear transfer from mature B and T donor cells. Nature 415:1035–1038
Humpherys D, Eggan K, Akutsu H, Hochedlinger K, Rideout WM 3rd, Biniszkiewicz D, Yanagimachi R, Jaenisch R (2001) Epigenetic instability in ES cells and cloned mice. Science 293:95–97
James RM, Kaufman MH, Webb S, West JD (1992) Electrofusion of mouse embryos results in uniform tetraploidy and not tetraploid/diploid mosaicism. Genet Res 60:185–194
Kaufman MH, Webb S (1990) Postimplantation development of tetraploid mouse embryos produced by electrofusion. Development 110:1121–1132
Li X, Yu Y, Wei W, Yong J, Yang J, You J, Xiong X, Qing T, Deng H (2005a) Simple and efficient production of mice derived from embryonic stem cells aggregated with tetraploid embryos. Mol Reprod Dev 71:154–158
Li X, Wei W, Yong J, Jia Q, Yu Y, Di K (2005b) The genetic heterozygosity and fitness of tetraploid embryos and embryonic stem cells are crucial parameters influencing survival of mice derived from embryonic stem cells by tetraploid embryo aggregation. Reproduction 130:53–59
Mann MR, Chung YG, Nolen LD, Verona RI, Latham KE, Bartolomei MS (2003) Disruption of imprinted gene methylation and expression in cloned preimplantation stage mouse embryos. Biol Reprod 69:902–914
McWhir J, Schnieke AE, Ansell R, Wallace H, Colman A, Scott AR, Kind AJ (1996) Selective ablation of differentiated cells permits isolation of embryonic stem cell lines from murine embryos with a non-permissive genetic background. Nat Genet 14:223–226
Nagy A, Gocza E, Diaz EM, Prideaux V, Ivçnyi E, Markkula M, Rossant J (1990) Embryonic stem cells alone are able to support fetal development in the mouse. Development 110:815–821
Nagy A, Rossant J, Nagy R, Abramow-Newerly W, Roder JC (1993) Derivation of completely cell culture-derived mice from early-passage embryonic stem cells. Proc Natl Acad Sci USA 90:8424–8428
Nagy A, Gertsenstein M, Vintersten K, Behringer R (2003) Manipulating the mouse embryo: a laboratory manual. Cold Spring Harbor, Cold Spring Harbor Laboratory Press, New York, pp 453–506
O’Shea KS (2004) Self-renewal vs. differentiation of mouse embryonic stem cells. Biol Reprod 71:1755–1765
Rachel E, Nissim B (2002) A molecular view on pluripotent stem cells. FEBS Lett 529:135–141
Rie S, Aya S, Michiko H, Akio T, Hiroyoshi I, Ying L, Satoshi K, Fumihiro S, Ken-ichi Y (2005) Establishment of a new embryonic stem cell line derived from C57BL/6 mouse expressing EGFP ubiquitously. Genesis 42:47–52
Schalkwyk LC, Jung M, Daser A, Weiher M, Walter J, Himmelbauer H, Lehrach H (1999) Panel of microsatellite markers for whole-genome scans and radiation hybrid mapping and a mouse family tree. Genome Res 9:878–887
Shiue YL, Liou JF, Shiau JW, Yang JR, Chen YH, Tailiu JJ, Chen LR (2006) In vitro culture period but not the passage number influences the capacity of chimera production of inner cell mass and its deriving cells from porcine embryos.Anim Reprod Sci 93:134–143
Schumacher A, Doerfler W (2004) Influence of in vitro manipulation on the stability of methylation patterns in the Snurf/Snrpn-imprinting region in mouse embryonic stem cells. Nucleic Acids Res 32:1566–1576
Schwenk F, Zevnik B, Bruning J, Rohl M, Willuweit A, Rode A, Hennek T, Kauselmann G, Jaenisch R, Kuhn R (2003) Hybrid embryonic stem cell-derived tetraploid mice show apparently normal morphological, physiological, and neurological characteristics. Mol Cell Biol 23:3982–3989
Seibler J, Zevnik B, Küter-Luks B, Andreas S, Kern H, Hennek T, Rode A, Heimann C, Faust N, Kauselmann G, Schoor M, Jaenisch R, Rajewsky K, Kühn R, Schwenk F (2003) Rapid generation of inducible mouse mutants. Nucleic Acids Res 31:e12
Wang Z, Jaenisch R (2004) At most three ES cells contribute to the somatic lineages of chimeric mice and of mice produced by ES-tetraploid complementation. Dev Biol 275:192–201
Wang ZQ, Kiefer F, Urbanek P, Wagner EF (1997) Generation of completely embryonic stem cell-derived mutant mice using tetraploid blastocyst injection. Mech Dev 62:137–145
Zhou D, Ren J, Ryan T, Higgins P, Tim M (2004) Rapid tagging of endogenous mouse genes by recombineering and ES cell complementation of tetraploid blastocysts. Nucleic Acids Res 32:e128
Zvetkova I, Apedaile A, Ramsahoye B, Mermoud JE, Crompton LA, John R, Feil R, Brockdorff N (2005) Global hypomethylation of the genome in XX embryonic stem cells. Nat Genet 37:1274–1279
We are grateful to Mrs. Jie Fang You and Mrs. Xiao Ran Xiong for culturing the ES cells used in our studies, to Ya Shu Liu for PCR primer pairs, and to Dr. Jie Yang for microsatellite DNA analysis.
This work was supported by the National Natural Science Foundation of China (grant no. 30571336) and the President Foundation of the Agricultural University of Hebei.
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Li, X., Jia, Q., Di, K. et al. Passage number affects the pluripotency of mouse embryonic stem cells as judged by tetraploid embryo aggregation. Cell Tissue Res 327, 607 (2007). https://doi.org/10.1007/s00441-006-0354-6
- Embryonic stem cells
- Tetraploid embryo