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Fibroblast cell line establishment, cryopreservation and interspecies embryos reconstruction in red panda (Ailurus fulgens)

Published online by Cambridge University Press:  01 May 2009

Yong Tao ,
Jianming Liu ,
Yunhai Zhang ,
Meiling Zhang ,
Junshun Fang ,
Wei Han ,
Zhizhong Zhang ,
Ya Liu ,
Jianping Ding  and
Xiaorong Zhang
Yong Tao
Affiliation:
College of Animal Science and Technology, Anhui Agricultural University, Hefei, 230036, China. These authors contributed equally to the paper.
Jianming Liu
Affiliation:
College of Animal Science and Technology, Anhui Agricultural University, Hefei, 230036, China. These authors contributed equally to the paper.
Yunhai Zhang
Affiliation:
College of Animal Science and Technology, Anhui Agricultural University, Hefei, 230036, China.
Meiling Zhang
Affiliation:
College of Animal Science and Technology, Anhui Agricultural University, Hefei, 230036, China.
Junshun Fang
Affiliation:
College of Animal Science and Technology, Anhui Agricultural University, Hefei, 230036, China.
Wei Han
Affiliation:
College of Animal Science and Technology, Anhui Agricultural University, Hefei, 230036, China.
Zhizhong Zhang
Affiliation:
Hefei Wild Animal Park, Hefei, 230061, Anhui Province, China.
Ya Liu
Affiliation:
College of Animal Science and Technology, Anhui Agricultural University, Hefei, 230036, China.
Jianping Ding
Affiliation:
College of Animal Science and Technology, Anhui Agricultural University, Hefei, 230036, China.
Xiaorong Zhang*
Affiliation:
College of Animal Science and Technology, Anhui Agricultural University, Changjiang West Road 130, Hefei, 230036, China.
*
All correspondence to: Zhang Xiaorong. College of Animal Science and Technology, Anhui Agricultural University, Changjiang West Road 130, Hefei, 230036, China. Tel: +8 6551 5782 488. Fax: +8 6551 5785 543. e-mail: [email protected]
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Summary

In evolution, the red panda (Ailurus fulgens) plays a pivotal role in the higher level phylogeny of arctoides carnivore mammals. The red panda inhabits certain Asian countries only and its numbers are decreasing. Therefore, the development of feasible ways to preserve this species is necessary. Genetic resource cryopreservation and somatic cell nuclear transfer (SCNT) have been used extensively to rescue this endangered species. The present study describes the establishment, for the first time, of a red panda ear fibroblast cell line, which was then cryopreserved, thawed and cultured. Through micromanipulation, interspecies embryos were reconstructed using the cryopreserved–thawed fibroblasts of the red panda as the donor and rabbit oocytes as recipients. A total of 194 enucleated rabbit oocytes were reconstructed with red panda ear fibroblasts; enucleated oocytes were activated without fusion as the control. The results show that the fibroblast cell line was established successfully by tissue culture and then cryopreserved in liquid nitrogen. Supplementation with 20% fetal bovine serum and 8% dimethyl sulphoxide in basic medium facilitated the cryopreservation. The interspecies embryos were successfully reconstructed. The cleavage, morulae and blastocyst rates after in vitro culture were 71, 47 and 23% (31/194), respectively. This study indicated that a somatic cell line could be established and cryopreserved from red panda and that rabbit cytoplast supports mitotic cleavage of the red panda karyoplasts and is capable of reprogramming the nucleus to achieve blastocysts.

Keywords

Ear fibroblastInterspecies somatic cell nuclear transferRabbitRed panda
Type
Research Article
Information
Zygote , Volume 17 , Issue 2 , May 2009 , pp. 117 - 124
Copyright
Copyright © Cambridge University Press 2009

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References

Borderie, V.M., Lopez, M., Lombet, A., Carvajal-Gonzalez, S., Cywiner, C. & Laroche, L. (1998). Cryopreservation and culture of human corneal keratocytes. Invest. Ophthalmol. Vis. Sci. 39, 1511–9.Google ScholarPubMed
Chen, D.Y., Sun, Q.Y., Liu, Y.L., Li, G.P., Lian, L., Wang, M.K., Han, M.Z., Song, X.F. & Sun, Q. (1999). The giant panda (Ailuropoda melanoleuca) somatic nucleus can dedifferentiate in rabbit ooplasm and support early development of the reconstructed egg. Sci. China (Ser C) 42, 346–53.Google Scholar
Choi, Y.H., Love, C.C., Chung, Y.G., Varner, D.D., Westhusin, M.E., Burghardt, R.C. & Hinrichs, K. (2002). Production of nuclear transfer horse embryos by piezo-driven injection of somatic cell nuclei and activation with stallion sperm cytosolic extract. Biol. Reprod. 67, 561–7.CrossRefGoogle ScholarPubMed
Dominko, T., Mitalipova, M., Haley, B., Beyhan, Z., Memili, E., McKusick, B. & First, N.L. (1999). Bovine oocyte cytoplasm supports development of embryos produced by nuclear transfer of somatic cell nuclei from various mammalian species. Biol. Reprod. 60, 1496–502.CrossRefGoogle ScholarPubMed
Flynn, J.J., Nedbal, M.A., Dragoo, J.W. & Honeycutt, R.L. (2000). Whence the red panda? Mol Phylogenet. Evo. 17, 190–9.CrossRefGoogle ScholarPubMed
Freshney, R.I. Disaggregation of the tissue and primary culture. (1994). In: Freshney, RI (ed.)., Culture of animal cells: a manual of basic technique. New York: Wiley-Liss & Sons., Inc. pp. 127–47.Google Scholar
Hayes, O., Rodriguez, L., Gonzalez, A., Falcon, V., Aguilar, A. & Castro, F.O. (2005). Effect of cryopreservation on fusion efficiency and in vitro development into blastocysts of bovine cell lines used in somatic cell cloning. Zygote 13, 277–82.CrossRefGoogle ScholarPubMed
Ikumi, S., Sawai, K., Takeuchi, Y., Iwayama, H., Ishikawa, H., Ohsumi, S. & Fukui, Y. (2004). Interspecies somatic cell nuclear transfer for in vitro production of Antarctic minke whale (Balaenoptera bonaerensis) embryos. Cloning Stem Cells 6, 284–93.CrossRefGoogle ScholarPubMed
Jiang, M.X., Yang, C.X., Zhang, L.S., Zheng, Y.L., Liu, S.Z., Sun, Q.Y. & Chen, D.Y. (2004). The effects of chemical enucleation combined with whole cell intracytoplasmic injection on panda–rabbit interspecies nuclear transfer. Zygote 12 (4), 315–20.Google Scholar
Kearney, J.N.Cryopreservation of cultured skin cells. (1991). Burns 1991., 17, 380–3.Google Scholar
Lanza, R.P., Cibelli, J.B., Diaz, F., Moraes, C.T., Farin, P.W., Farin, C.E., Hammer, C.J., West, M.D. & Damiani, P. (2000). Cloning of an endangered species (Bos gaurus) using interspecies nuclear transfer. Cloning 2, 79–90.CrossRefGoogle ScholarPubMed
Lee, B., Wirtu, G.G., Damiani, P., Pope, E., Dresser, B.L., Hwang, W. & Bavister, B.D. (2003). Blastocyst development after intergeneric nuclear transfer of mountain bongo antelope somatic cells into bovine oocytes. Cloning Stem Cells 5, 25–33.CrossRefGoogle ScholarPubMed
Li, J.S., Han, Z.M., Zhu, Z.Y., Wen, D.C., Liu, Z.H., Wang, M.K., Lian, L., Du, J., Wang, P.Y., Zhang, H.M. & Chen, D.Y. (2002). Interspecies nuclear transfer using nonquiescent somatic cell. J. Exp. Biol. 35, 62–5.Google Scholar
Li, M., Wei, F., Goossens, B., Feng, Z., Tamate, H.B., Bruford, M.W. & Funk, S.M. (2005). Mitochondrial phylogeography and subspecific variation in the red panda (Ailurus fulgens): implications for conservation. Mol. Phylogenet. Evol. 36, 78–89.CrossRefGoogle ScholarPubMed
Liu, S.Z., Zhou, Z.M., Chen, T., Zhang, Y.L., Wen, D.C., Kou, Z.H., Li, Z.D., Sun, Q.Y. & Chen, D.Y. (2004). Blastocysts produced by nuclear transfer between chicken blastodermal cells and rabbit oocytes. Mol. Reprod. Dev. 69, 296–302.CrossRefGoogle ScholarPubMed
Liu, Y., Zhang, X.R., Chen, D.Y., Zhang, Y.H., Zhang, Z.G., Jin, R.T., Wang, C.L., Zhang, M.L., Li, D.W., Li, B., Zhao, H. & Cheng, L.Z. (2003). In vitro developmental potential of cloned embryos derived from bovine somatic and rabbit oocyte. Agric. Sci. China 2, 1393–7.Google Scholar
Liu, Y., Zhang, X.R., Zhang, Y.H., Jin, R.T., Wang, C.L. & Wu, J.Y. (2004a). Storage and cryopreservation of bovine and goat fibroblast (in Chinese). Chin. J. Vet. Sci. 24, 197–8.Google Scholar
Liu, Y., Zhang, X.R., Chen, D.Y., Zhang, Y.H., Zhang, Z.G., Jin, R.T., Wang, C.L., Zhang, M.L., Li, D.W., Lin, B., Zhao, H. & Cheng, L.Z. (2004b). Study on development of cloned embryo using bovine somatic cell and rabbit oocyte in vitro. Scientia Agricultura Sinica 2004b; 37, 441–5.Google Scholar
Loi, P., Ptak, G., Barboni, B., Fulka, J., Cappai, P. & Clinton, M. (2001). Genetic rescue of an endangered mammal by cross species nuclear transfer using post-mortem somatic cells. Nature Biotech. 19, 962–4.CrossRefGoogle ScholarPubMed
Lu, F., Shi, D; Wei, J., Yang, S. & Wei, Y. (2005). Development of embryos reconstructed by interspecies nuclear transfer of adult fibroblasts between buffalo (Bubalus bubalis) and cattle (Bos indicus). Theriogenology 64, 1309–19.CrossRefGoogle ScholarPubMed
Murakami, M., Otoi, T., Wongsrikeao, P., Agung, B., Sambuu, R. & Suzuki, T. (2005). Development of interspecies cloned embryos in yak and dog. Cloning Stem Cells 7, 77–81.CrossRefGoogle ScholarPubMed
Peura, T.T. (2000). Serum starvation can cause excessive DNA damage in sheep fetal fibroblasts. Theriogenology 55, 285 (Abstract).Google Scholar
Peura, T.T., Hartwich, K.M., Hamilton, H.M. & Walker, S.K. (2003). No differences in sheep somatic cell nuclear transfer outcomes using serum-starved or actively growing donor granulosa cells. Reprod. Fertil. Dev. 15, 157–65.CrossRefGoogle ScholarPubMed
Sansinena, M.J., Hylan, D., Hebert, K., Denniston, R.S. & Godke, R.A. (2005). Banteng (Bos javanikus) embryos and pregnancies produced by interspecies nuclear transfer. Theriogenology 63, 1081–91.Google Scholar
Slattery, J.P. & O'Brien, S.J. (1995). Molecular phylogeny of the red panda (Ailurus fulgens). J. Hered. 86, 413–22.Google Scholar
Tani, T., Kate, K. & Tsunoda, Y. (2000). Developmental potential of cumulus cell-derived culture frozen in a quiescent state nuclear transfer. Theriogenology 53, 1623–9.CrossRefGoogle Scholar
Thongphakdee, A., Numchaisrika, P., Omsongkram, S., Chatdarong, K., Kamolnorranath, S., Dumnui, S. & Techakumphu, M. (2006). In vitro development of marbled cat embryos derived from interspecies somatic cell nuclear transfer. Reprod. Dom. Anim. 41, 219–26.CrossRefGoogle ScholarPubMed
Tian, Y., Nie, W.H., Wang, J.H., Yang, Y.F. & Yang, F.T. (2002). Comparative chromosome painting shows the red panda (Ailurus fulgens) has a highly conserved karyotype. Acta Genetic Sinica 29, 124–7.Google Scholar
Wen, D.C., Yang, C.X., Cheng, Y., Li, J.S., Liu, Z.H., Sun, Q.Y., Zhang, J.X., Lei, L., Wu, Y.Q., Kou, Z.H. & Chen, D.Y. (2003). Comparison of developmental capacity for intra- and interspecies cloned cat (Felis catus) embryos. Mol. Reprod. Dev. 66, 38–45.CrossRefGoogle ScholarPubMed
Wen, D.C., Bi, C.M. & Chen, D.Y. (2004). Progress in interspecies cloning of mammals. Prog. Natur. Sci. 14, 1–8.Google Scholar
Wen, D.C., Bi, C.M., Xu, Y., Yang, C.X., Zhu, Z.Y., Sun, Q.Y. & Chen, D.Y. (2005). Hybrid embryos produced by transferring panda or cat somatic nuclei into rabbit MII oocytes can develop to blastocyst in vitro. J. Exp. Zool. 303, 689–97.CrossRefGoogle ScholarPubMed
White, K.L., Bunch, T.D., Mitalipov, S. & Reed, W.A. (1999). Establishment of pregnancy after the transfer of nuclear transfer embryos produced from the fusion of argali (Ovis ammon) nuclei into domestic sheep (Ovis aries) enucleated oocytes. Cloning 1, 47–54.Google Scholar
Zhang, X.R., Liu, Y., Zhang, Y.H., Chen, D.Y., Lian, L., Li, J.S., Jin, R.T., Zhang, Z.G., Zhang, M.L. & Wang, C.L. (2004). Effect of some factors on the fusion rate of bovine–rabbit interspecies reconstructed eggs. Chin. J. Agric. Biotech. 1, 135–8.Google Scholar
Zhang, Y.H., Pan, D.K., Sun, X.Z., Sun, G.J., Wang, X.B., Liu, X.H., Li, Y., Dai, Y.P. & Li, N. (2006). Production of porcine cloned transgenic embryos expressing green fluorescent protein by somatic cell nuclear transfer. Sci. Chin. (Series C) 149, 1–11.Google Scholar
Zhang, Z.G., Zhang, X.R., Liu, Y., Jin, R.T., Zhang, M.L., Wang, C.L. & Zhao, H. (2005). Study of serial nuclear transfer on goat (Bore) rabbit interspecies cloned embryo. Scientia Agricultura Sinica 38, 601–5.Google Scholar
Zhong, Z.S., Zhang, G., Meng, X.Q., Zhang, Y.L., Chen, D.Y., Schatten, H. & Sun, Q.Y. (2005). Function of donor cell centrosome in intraspecies and interspecies nuclear transfer embryos. Exp. Cell Res. 306, 35–46.CrossRefGoogle ScholarPubMed