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Cell-cycle synchronization of fibroblasts derived from transgenic cloned cattle ear skin: effects of serum starvation, roscovitine and contact inhibition

Published online by Cambridge University Press:  01 May 2008

XiuZhu Sun ,
ShuHui Wang ,
YunHai Zhang ,
HaiPing Wang ,
LiLi Wang ,
Liu Ying ,
Rong Li  and
Ning Li
XiuZhu Sun
Affiliation:
State Key Laboratory for Agrobiotechnology, China Agricultural University, Beijing 100094, China.
ShuHui Wang
Affiliation:
Institute of Animal Science Chinese Academy of Agricultural Science Beijing100094, China.
YunHai Zhang
Affiliation:
AnHui Agricultural University, HeFei, 230036, AnHui province, China.
HaiPing Wang
Affiliation:
Beijing GenProtein Biotechnology Ltd, BeiJing, 100094, China.
LiLi Wang
Affiliation:
Beijing GenProtein Biotechnology Ltd, BeiJing, 100094, China.
Liu Ying
Affiliation:
Beijing GenProtein Biotechnology Ltd, BeiJing, 100094, China.
Rong Li
Affiliation:
Beijing GenProtein Biotechnology Ltd, BeiJing, 100094, China.
Ning Li*
Affiliation:
State Key Laboratory for Agrobiotechnology, College of Biological Science China Agricultural University, Beijing 100094, China. State Key Laboratory for Agrobiotechnology, China Agricultural University, Beijing 100094, China.
*
All correspondence to: Ning Li. State Key Laboratory for Agrobiotechnology, College of Biological Science China Agricultural University, Beijing 100094, China. Tel: +86 10 62733323. Fax: +86 10 62733904. e-mail: [email protected]
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Summary

The purpose of the present study was to evaluate the effects of serum-starvation, contact-inhibition and roscovitine treatments on cell-cycle synchronization at the G0/G1 stage of ear skin fibroblasts isolated from transgenic cloned cattle. The developmental competence of re-cloned embryos was also examined. Our results showed that the proportion of G0/G1 cells from the serum-starved group at 3, 4 or 5 days was significantly higher compared with 1 or 2 days only (91.5, 91.7 and 93.5% versus 90.1 and 88.8%, respectively, p < 0.05); whilst there was no statistical difference among cells at 3, 4 or 5 days. For roscovitine-treated cells, the proportion of G0/G1 cells at 2, 3, 4 or 5 days was significantly higher than those treated for 1 day only (91.1, 90.1, 89.4 and 91.3% versus 86.51%, respectively, p < 0.05). The proportion of contact-inhibited G0/G1 cells rose significantly with treatment time, but was similar at 3, 4 and 5 days (89.4, 90.4, 91.4, 91.6 and 92.1%, respectively, p < 0.05). The efficiency of obtaining G0/G1 phase cells was lower when roscovitine treatment was employed to synchronize the cell cycle compared with the serum-starvation and contact-inhibition methods (89.7 versus 91.1% and 91.0%, p < 0.05). Moreover, obvious differences were observed in the rate of fused couplets and blastocysts (89.88 ± 2.70 versus 87.40 ± 5.13; 44.10 ± 8.62 versus 58.38 ± 13.28, respectively, p < 0.05), when nuclear transfer embryos were reconstructed using donors cells that had been serum starved or contact inhibited for 3 days. Our data indicate that 3 day treatment is feasible for harvesting sufficient G0/G1 cells to produce re-cloned transgenic bovine embryos, regardless of whether serum-starvation, contact-inhibition or roscovitine treatments are used as the synchronization methods.

Keywords

Cell-cycle synchronizationFlow cytometry Somatic cell nuclear transferTransgenic
Type
Research Article
Information
Zygote , Volume 16 , Issue 2 , May 2008 , pp. 111 - 116
Copyright
Copyright © Cambridge University Press 2008

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References

Alessi, F., Quarta, S., Savio, M., Riva, F., Rossi, L., Stivala, L.A., Scovassi, A.I., Meijer, L. & Prosperi, E. (1998). The cyclin-dependent kinase inhibitors olomoucine and roscovitine arrest human fibroblasts in G1 phase by specific inhibition of CDK2 kinase activity. Exp. Cell. Res. 245, 8–18.CrossRefGoogle ScholarPubMed
Baguisi, A., Behboodi, E., Melican, D.T., Pollock, J.S., Destrempes, M.M., Cammuso, C., Williams, J.L., Nims, S.D., Porter, C.A., Midura, P., Palacios, M.J., Ayres, S.L., Denniston, R.S., Hayes, M.L., Ziomek, C.A., Meade, H.M., Godke, R.A., Gavin, W.G., Overstrom, E.W. & Echelard, Y. (1999). Production of goats by somatic cell nuclear transfer. Nat. Biotechnol. 17, 456–61.CrossRefGoogle ScholarPubMed
Boquest, A.C., Day, B.N. & Prather, R.S. (1999). Flow cytometric cell cycle analysis of cultured porcine fetal fibroblast cells. Biol. Reprod. 60, 1013–9.CrossRefGoogle ScholarPubMed
Campbell, K.H., Loi, P., Otaegui, P.J. & Wilmut, I. (1996). Cell cycle co-ordination in embryo cloning by nuclear transfer. Rev. Reprod. 1, 40–6.CrossRefGoogle ScholarPubMed
Chesne, P., Adenot, P.G., Viglietta, C., Baratte, M., Boulanger, L. & Renard, J.P. (2002). Cloned rabbits produced by nuclear transfer from adult somatic cells. Nat. Biotechnol. 20, 366–9.CrossRefGoogle ScholarPubMed
Cho, S.R., Ock, S.A., Yoo, J.G., Mohana Kumar, B., Choe, S.Y. & Rho, G.J. (2005). Effects of confluent, roscovitine treatment and serum starvation on the cell-cycle synchronization of bovine foetal fibroblasts. Reprod. Domest. Anim. 40, 171–6.CrossRefGoogle Scholar
Cibelli, J.B., Stice, S.L., Golueke, P.J., Kane, J.J., Jerry, J., Blackwell, C., Ponce de Leon, F.A. & Robl, J.M. (1998). Transgenic bovine chimeric offspring produced from somatic cell-derived stem-like cells. Nat. Biotechnol. 16, 642–6.CrossRefGoogle ScholarPubMed
Galli, C., Lagutina, I., Crotti, G., Colleoni, S., Turini, P., Ponderato, N., Duchi, R. & Lazzari, G. (2003). Pregnancy: a cloned horse born to its dam twin. Nature, 424, 635.CrossRefGoogle ScholarPubMed
Gibbons, J., Arat, S., Rzucidlo, J., Miyoshi, K., Waltenburg, R., Respess, D., Venable, A. & Stice, S. (2002). Enhanced survivability of cloned calves derived from roscovitine-treated adult somatic cells. Biol. Reprod. 66, 895–900.CrossRefGoogle ScholarPubMed
Holden, C. (2003). Genetics. First cloned mule races to finish line. Science 300, 1354.CrossRefGoogle ScholarPubMed
Kasinathan, P., Knott, J.G., Moreira, P.N., Burnside, A.S., Jerry, D.J. & Robl, J.M. (2001). Effect of fibroblast donor cell age and cell cycle on development of bovine nuclear transfer embryos in vitro. Biol. Reprod. 64, 1487–93.CrossRefGoogle ScholarPubMed
Kato, Y., Tani, T., Sotomaru, Y., Kurokawa, K., Kato, J., Doguchi, H., Yasue, H. & Tsunoda, Y. (1998). Eight calves cloned from somatic cells of a single adult. Science 282, 2095–8.CrossRefGoogle ScholarPubMed
Kubota, C., Yamakuchi, H., Todoroki, J., Mizoshita, K., Tabara, N., Barber, M. & Yang, X. (2000). Six cloned calves produced from adult fibroblast cells after long-term culture. Proc. Natl. Acad. Sci. USA 97, 990–5.CrossRefGoogle ScholarPubMed
Lai, L., Park, K.W., Cheong, H.T., Kuhholzer, B., Samuel, M., Bonk, A., Im, G.S., Rieke, A., Day, B.N., Murphy, C.N., Carter, D.B. & Prather, R.S. (2002). Transgenic pig expressing the enhanced green fluorescent protein produced by nuclear transfer using colchicine-treated fibroblasts as donor cells. Mol. Reprod. Dev. 62, 300–6.CrossRefGoogle ScholarPubMed
Liu, C.T., Yu, K.C. & Ju, J.C. (2004). Cell cycle stage analysis of rabbit foetal fibroblasts and cumulus cells. Reprod. Domest. Anim. 39, 385–90.CrossRefGoogle ScholarPubMed
Polejaeva, I.A., Chen, S.H., Vaught, T.D., Page, R.L., Mullins, J., Ball, S., Dai, Y., Boone, J., Walker, S., Ayares, D.L., Colman, A. & Campbell, K.H. (2000). Cloned pigs produced by nuclear transfer from adult somatic cells. Nature 407, 86–90.CrossRefGoogle ScholarPubMed
Prather, R.S., Boquest, A.C. & Day, B.N. (1999). Cell cycle analysis of cultured porcine mammary cells. Cloning 1, 17–24.CrossRefGoogle ScholarPubMed
Qi, Z., Kishigami, A., Nakagawa, Y., Iida, H. & Sokabe, M. (2004). A mechanosensitive anion channel in Arabidopsis thaliana mesophyll cells. Plant Cell Physiol. 45, 1704–8.CrossRefGoogle ScholarPubMed
Renard, J.P., Zhou, Q., LeBourhis, D., Chavatte-Palmer, P., Hue, I., Heyman, Y. & Vignon, X. (2002). Nuclear transfer technologies: between successes and doubts. Theriogenology 57, 203–22.CrossRefGoogle ScholarPubMed
Shin, T., Kraemer, D., Pryor, J., Liu, L., Rugila, J., Howe, L., Buck, S., Murphy, K., Lyons, L. & Westhusin, M. (2002). A cat cloned by nuclear transplantation. Nature 415, 859.CrossRefGoogle ScholarPubMed
Urakawa, M., Ideta, A., Sawada, T. & Aoyagi, Y. (2004). Examination of a modified cell cycle synchronization method and bovine nuclear transfer using synchronized early G1 phase fibroblast cells. Theriogenology 62, 714–28.CrossRefGoogle ScholarPubMed
Wakayama, T., Perry, A.C., Zuccotti, M., Johnson, K.R. & Yanagimachi, R. (1998). Full-term development of mice from enucleated oocytes injected with cumulus cell nuclei. Nature 394, 369–74.CrossRefGoogle ScholarPubMed
Wells, D.N., Misica, P.M. & Tervit, H.R. (1999). Production of cloned calves following nuclear transfer with cultured adult mural granulosa cells. Biol. Reprod. 60, 996–1005.CrossRefGoogle ScholarPubMed
Wilmut, I., Schnieke, A.E., McWhir, J., Kind, A.J. & Campbell, K.H. (1997). Viable offspring derived from fetal and adult mammalian cells. Nature 385, 810–3.CrossRefGoogle ScholarPubMed
Yin, X.J., Lee, H.S., Kim, L.H., Shin, H.D., Kim, N.H. & Kong, I.K. (2007). Effect of serum starvation on the efficiency of nuclear transfer using odd-eyed white cat fibroblasts. Theriogenology 67, 816–23.CrossRefGoogle ScholarPubMed
Zakhartchenko, V., Durcova-Hills, G., Schernthaner, W., Stojkovic, M., Reichenbach, H.D., Mueller, S., Steinborn, R., Mueller, M., Wenigerkind, H., Prelle, K., Wolf, E. & Brem, G. (1999). Potential of fetal germ cells for nuclear transfer in cattle. Mol. Reprod. Dev. 52, 421–6.3.0.CO;2-P>CrossRefGoogle ScholarPubMed
Zetterberg, A. & Auer, G. (1970). Proliferative activity and cytochemical properties of nuclear chromatin related to local cell density of epithelial cells. Exp. Cell Res. 62, 262–70.CrossRefGoogle ScholarPubMed
Zhou, Q., Jouneau, A., Brochard, V., Adenot, P. & Renard, J. P. (2001). Developmental potential of mouse embryos reconstructed from metaphase embryonic stem cell nuclei. Biol. Reprod. 65, 412–9.CrossRefGoogle ScholarPubMed
Zhou, Q., Yang, S.H., Ding, C.H., He, X.C., Xie, Y.H., Hildebrandt, T.B., Mitalipov, S.M., Tang, X.H., Wolf, D.P. & Ji, W.Z. (2006). A comparative approach to somatic cell nuclear transfer in the rhesus monkey. Hum. Reprod. 21, 2564–71.CrossRefGoogle ScholarPubMed