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Journal of Zhejiang University SCIENCE B 2007 Vol.8 No.2 P.136-146


Ex vivo expansion and pluripotential differentiation of cryopreserved human bone marrow mesenchymal stem cells

Author(s):  XIANG Ying, ZHENG Qiang, JIA Bing-bing, HUANG Guo-ping, Xu Yu-lin, WANG Jin-fu, PAN Zhi-jun

Affiliation(s):  School of Life Sciences, Zhejiang University, Hangzhou 310012, China; more

Corresponding email(s):   wjfu@zju.edu.cn, zepzj@163.com

Key Words:  Bone marrow mesenchymal stem cells, Cryopreservation, Expansion, Differentiation

XIANG Ying, ZHENG Qiang, JIA Bing-bing, HUANG Guo-ping, Xu Yu-lin, WANG Jin-fu, PAN Zhi-jun. Ex vivo expansion and pluripotential differentiation of cryopreserved human bone marrow mesenchymal stem cells[J]. Journal of Zhejiang University Science B, 2007, 8(2): 136-146.

@article{title="Ex vivo expansion and pluripotential differentiation of cryopreserved human bone marrow mesenchymal stem cells",
author="XIANG Ying, ZHENG Qiang, JIA Bing-bing, HUANG Guo-ping, Xu Yu-lin, WANG Jin-fu, PAN Zhi-jun",
journal="Journal of Zhejiang University Science B",
publisher="Zhejiang University Press & Springer",

%0 Journal Article
%T Ex vivo expansion and pluripotential differentiation of cryopreserved human bone marrow mesenchymal stem cells
%A ZHENG Qiang
%A JIA Bing-bing
%A HUANG Guo-ping
%A Xu Yu-lin
%A WANG Jin-fu
%A PAN Zhi-jun
%J Journal of Zhejiang University SCIENCE B
%V 8
%N 2
%P 136-146
%@ 1673-1581
%D 2007
%I Zhejiang University Press & Springer
%DOI 10.1631/jzus.2007.B0136

T1 - Ex vivo expansion and pluripotential differentiation of cryopreserved human bone marrow mesenchymal stem cells
A1 - XIANG Ying
A1 - ZHENG Qiang
A1 - JIA Bing-bing
A1 - HUANG Guo-ping
A1 - Xu Yu-lin
A1 - WANG Jin-fu
A1 - PAN Zhi-jun
J0 - Journal of Zhejiang University Science B
VL - 8
IS - 2
SP - 136
EP - 146
%@ 1673-1581
Y1 - 2007
PB - Zhejiang University Press & Springer
ER -
DOI - 10.1631/jzus.2007.B0136

This study is aimed at investigating the potentials of ex vivo expansion and pluri-differentiation of cryopreservation of adult human bone marrow mesenchymal stem cells (hMSCs) into chondrocytes, adipocytes and neurocytes. Cryopreserved hMSCs were resuscitated and cultured for 15 passages, and then induced into chondrocytes, adipocytes and neurocytes with corresponding induction medium. The induced cells were observed for morphological properties and detected for expressions of type II collagen, triglyceride or neuron-specific enolase and nestin. The result showed that the resuscitated cells could differentiate into chondrocytes after exposure to transforming growth factor β1 (TGF-β1), insulin-like growth factor I (IGF-I) and vitamin C (VC), and uniformly changed morphologically from a spindle-like fibroblastic appearance to a polygonal shape in three weeks. The induced cells were heterochromatic to safranin O and expressed cartilage matrix-procollagenal (II) mRNA. The resuscitated cells cultured in induction medium consisting of dexamethasone, 3-isobutyl-1-methylxanthine, indomethacin and IGF-I showed adipogenesis, and lipid vacuoles accumulation was detectable after 21 d. The resuscitated hMSCs were also induced into neurocytes and expressed nestin and neuron specific endolase (NSE) that were special surface markers associated with neural cells at different stage. This study suggested that the resuscitated hMSCs should be still a population of pluripotential cells and that it could be used for establishing an abundant hMSC reservoir for further experiment and treatment of various clinical diseases.

Darkslateblue:Affiliate; Royal Blue:Author; Turquoise:Article


[1] Bai, X., Xiao, Z., Pan, Y., Hu, J., Pohl, J., Wen, J., Li, L., 2004. Cartilage-derived morphogenetic protein-1 promotes the differentiation of mesenchymal stem cells into chondrocytes. Biochem. Biophys. Res. Commun., 325(2):453-460.

[2] Bainter, J.J., Boos, A., Kroll, K.L., 2001. Neural induction takes a transcriptional twist. Dev. Dyn., 222(3):315-327.

[3] Blondheim, N.R., Levy, Y.S., Zur, T.B., Burshtein, A., Cherlow, T., Kan, I., Barzilai, R., Bahat-Stromza, M., Barhum, Y., Bulvik, S., et al., 2006. Human mesenchymal stem cells express neural genes, suggesting a neural predisposition. Stem Cells Dev., 15(2):141-164.

[4] Bruder, S.P., Jaiswal, N., Haynesworth, S.E., 1997. Growth kinetics, self-renewal, and the osteogenic potential of purified human mesenchymal stem cells during extensive subcultivation and following cryopreservation. J. Cell. Biochem., 64(2):278-294.

[5] Chen, F., Mao, T., Ding, G., 2003. Experimental study on the chondrogenesis potentiality of marrow stromal cell under the induction of transforming growth factor-β. West China J. Stomoatol., 21(2):92-94 (in Chinese).

[6] Cho, K.J., Trzaska, K.A., Greco, S.J., McArdle, J., Wang, F.S., Ye, J.H., Rameshwar, P., 2005. Neurons derived from human mesenchymal stem cells show synaptic transmission and can be induced to produce the neurotransmitter substance P by interleukin-1α. Stem Cells, 23(3):383-391.

[7] Chu, Q., Wang, Y., Fu, X., Zhang, S., 2004. Mechanism of in vitro differentiation of bone marrow stromal cells into neuron-like cells. J. Huazhong Univ. Sci. Technol. Med. Sci., 24(3):259-261.

[8] Collas, P., Hakelien, A.M., 2003. Teaching cells new tricks. Trends Biotechnol., 21(8):354-361.

[9] Dezawa, M., Kanno, H., Hoshino, M., Cho, H., Matsumoto, N., Itokazu, Y., Tajima, N., Yamada, H., Sawada, H., Ishikawa, H., et al., 2004. Specific induction of neuronal cells from bone marrow stromal cells and application for autologous transplantation. J. Clin. Invest., 113(12):1701-1710.

[10] Digirolamo, C.M., Stokes, D., Colter, D., Phinney, D.G., Class, R., Prockop, D.J., 1999. Propagation and senescence of human marrow stromal cells in culture: a simple colony-forming assay identifies samples with the greatest potential to propagate and differentiate. Br. J. Haematol., 107(2):275-281.

[11] Guo, Z.K., Liu, X.D., Hou, C.M., Li, X.S., Mao, N., 2001. Human bone marrow mesenchymal stem cells differentiate into neuron-like cells in vitro. J. Exp. Hematol. Sinica, 9(1):91-92 (in Chinese).

[12] Hayflick, L., 1989. Antecedents of aging research. Exp. Gerontol., 24(5-6):355-365.

[13] Indrawattana, N., Chen, G., Tadokoro, M., Shann, L.H., Ohgushi, H., Tateishi, T., Tanaka, J., Bunyaratvej, A., 2004. Growth factor combination for chondrogenic induction from human mesenchymal stem cell. Biochem. Biophys. Res. Commun., 320(3):914-919.

[14] Jiang, Y., Jahagirdar, B.N., Reinhardt, R.L., Schwartz, R.E., Keene, C.D., Ortiz-Gonzalez, X.R., Reyes, M., Lenvik, T., Lund, T., Blackstad, M., et al., 2002. Pluripotency of mesenchymal stem cells derived from adult marrow. Nature, 418(6893):41-49.

[15] Johnstone, B., Hering, T.M., Caplan, A.I., Goldberg, V.M., Yoo, J.U., 1998. In vitro chondrogenesis of bone marrow-derived mesenchymal progenitor cells. Exp. Cell Res., 238(1):265-272.

[16] Jori, F.P., Napolitano, M.A., Melone, M.A., Cipollaro, M., Cascino, A., Altucci, L., Peluso, G., Giordano, A., Galderisi, U., 2005. Molecular pathways involved in neural in vitro differentiation of marrow stromal stem cells. J. Cell. Biochem., 94(4):645-655.

[17] Kleber, M., Lee, H.Y., Wurdak, H., Buchstaller, J., Riccomagno, M.M., Ittner, L.M., Suter, U., Epstein, D.J., Sommer, L., 2005. Neural crest stem cell maintenance by combinatorial Wnt and BMP signaling. J. Cell Biol., 169(2):309-320.

[18] Korbling, M., Estrov, Z., 2003. Adult stem cells for tissue repair—a new therapeutic concept? N. Engl. J. Med., 349(6):570-582.

[19] Kotobuki, N., Hirose, M., Takakura, Y., Ohgushi, H., 2004. Cultured autologous human cells for hard tissue regeneration: preparation and characterization of mesenchymal stem cells from bone marrow. Artif. Organs, 28(1):33-39.

[20] Kotobuki, N., Hirose, M., Machida, H., Katou, Y., Muraki, K., Takakura, Y., Ohgushi, H., 2005. Viability and osteogenic potential of cryopreserved human bone marrow-derived mesenchymal cells. Tissue Eng., 11(5-6):663-673.

[21] Lee, J.W., Kim, Y.H., Kim, S.H., Han, S.H., Hahn, S.B., 2004. Chondrogenic differentiation of mesenchymal stem cells and its clinical applications. Yonsei Med. J., 45(Suppl.):41-47.

[22] Lisignoli, G., Cristino, S., Piacentini, A., Toneguzzi, S., Grassi, F., Cavallo, C., Zini, N., Solimando, L., Maraldi, N.M., Facchini, A., 2005. Cellular and molecular events during chondrogenesis of human mesenchymal stromal cells grown in a three-dimensional hyaluronan based scaffold. Biomaterials, 26(28):5677-5686.

[23] Lu, P., Blesch, A., Tuszynski, M.H., 2004. Induction of bone marrow stromal cells to neurons: differentiation, transdifferentiation, or artifact? J. Neurosci. Res., 77(2):174-191.

[24] Mackay, A.M., Beck, S.C., Murphy, J.M., Barry, F.P., Chichester, C.O., Pittenger, M.F., 1998. Chondrogenic differentiation of cultured human mesenchymal stem cells from marrow. Tissue Eng., 4(4):415-428.

[25] Meisler, M.H., Howard, G., 1989. Effects of insulin on gene transcription. Ann. Rev. Physiol., 51(1):701-714.

[26] Munoz-Elias, G., Woodbury, D., Black, I.B., 2003. Marrow stromal cells, mitosis, and neuronal differentiation: stem cell and precursor functions. Stem Cells, 21(4):437-448.

[27] Neuhuber, B., Gallo, G., Howard, L., Kostura, L., Mackay, A., Fisher, I., 2004. Reevaluation of in vitro differentiation protocols for bone marrow stromal cells: disruption of actin cytoskeleton induces rapid morphological changes and mimics neuronal phenotype. J. Neurosci. Res., 77(2):192-204.

[28] Otero, J.J., Fu, W., Kan, L., Cuadra, A.E., Kessler, J.A., 2004. β-catenin signaling is required for neural differentiation of embryonic stem cells. Development, 131(15):3545-3557.

[29] Pittenger, M.F., Mackay, A.M., Beck, S.C., Jaiswal, R.K., Douglas, R., Mosca, J.D., Moorman, M.A., Simonetti, D.W., Craig, S., Marshak, D.R., 1999. Multilineage potential of adult human mesenchymal stem cells. Science, 284(5411):143-147.

[30] Poh, M., Boyer, M., Solan, A., Dahl, S.L.M., Pedrotty, D., Banik, S.S.R., McKee, J.A., Klinger, R.Y., Counter, C.M., Niklason, L.E., 2005. Blood vessels engineered from human cells. Lancet, 365(9477):2122-2124.

[31] Qiu, L.Y., Wang, J.F., Shen, D., Jin, J., 2004. Expansion and chondrogenic induction of human bone marrow mesenchymal stem cells. J. Zhejiang Univ. (Sci. Ed.), 31(2):337-342 (in Chinese).

[32] Ren, T., Cao, Y., Zhao, Q., Zhou, C., Liao, L., Jia, M., Zhao, Q., Cai, H., Han, Z.C., Yang, R., et al., 2006. Proliferation and differentiation of bone marrow stromal cells under hypoxic conditions. Biochem. Biophys. Res. Commun., 347(1):12-21.

[33] Sanchez-Ramos, J., Song, S., Cardozo-Pelaez, F., Hazzi, C., Stedeford, T., Willing, A., Freeman, T.B., Saporta, S., Janssen, W., Patel, N., et al., 2000. Adult bone marrow stromal cells differentiate into neural cells in vitro. Exp. Neurol., 164(2):247-256.

[34] Shi, S., Gronthos, S., Chen, S., Reddi, A., Counter, C.M., Robey, P.G., Wang, C.Y., 2002. Bone formation by human postnatal bone marrow stromal stem cells is enhanced by telomerase expression. Nat. Biotechnol., 20(6):587-591.

[35] Simonsen, J.L., Rosada, C., Serakinci, N., Justesen, J., Stenderup, K., Rattan, S.I., Jensen, T.G., Kassem, M., 2002. Telomerase expression extends the proliferative life-span and maintains the osteogenic potential of human bone marrow stromal cells. Nat. Biotechnol., 20(6):592-596.

[36] Suon, S., Jin, H., Donaldson, A.E., Caterson, E.J., Tuan, R.S., Deschennes, G., Marshall, C., Iacovitti, L., 2004. Transient differentiation of adult human bone marrow cells into neuron-like cells in culture: development of morphological and biochemical traits is mediated by different molecular mechanisms. Stem Cells Dev., 13(6):625-635.

[37] Tondreau, T., Meuleman, N., Delforge, A., Dejeneffe, M., Leroy, R., Massy, M., Mortier, C., Bron, D., Lagneaux, L., 2005. Mesenchymal stem cells derived from CD133-positive cells in mobilized peripheral blood and cord blood: proliferation, Oct4 expression, and plasticity. Stem Cells, 23(8):1105-1112.

[38] Wang, J., Luo, C.J., Guo, C.H., Zhang, Y., 2002. Mesenchymal stem cells and related factors. J. Exp. Hematol. Sinica, 10(5):468-471 (in Chinese).

[39] Watanabe, H., de Caestecker, M.P., Yamada, Y., 2001. Transcriptional cross-talk between Smad, ERK1/2, and p38 mitogen-activated protein kinase pathways regulates transforming growth factor-β-induced aggrecan gene expression in chondrogenic ATDC5 cells. J. Biol. Chem., 276(17):14466-14473.

[40] Woodbury, D., Schwarz, E.J., Prockop, D.J., Black, I.B., 2000. Adult rat and human bone marrow stromal cells differentiate into neurons. J. Neurosci. Res., 61(4):364-370.

[41] Worster, A.A., Brower-Toland, B.D., Fortier, L.A., Bent, S.J., Williams, J., Nixon, A.J., 2001. Chondrocytic differentiation of mesenchymal stem cells sequentially exposed to transforming growth factor-β1 in monolayer and insulin-like growth factor-I in a three-dimensional matrix. J. Orthop. Res., 19(4):738-749.

[42] Xiang, P., Xia, W.J., Zhang, L.R., Chen, Z.G., Zhang, X.M., Li, Y., Li, S.N., 2001a. Human mesenchymal stem cells differentiate into neuron-like cells. Chin. J. Pathophysiol., 17(4):385-387 (in Chinese).

[43] Xiang, P., Zhang, L.R., Chen, Z.G., Xia, W.J., Zhang, X.M., Li, Y., Li, S.N., 2001b. Adult human mesenchymal stem cells differentiate into adipocytes. Chin. J. Pathophysiol., 17(6):598-601 (in Chinese).

[44] Yamaguchi, M., Hirayama, F., Kanai, M., Sato, N., Fukazawa, K., Yamashita, K., Sawada, K., Koike, T., Kuwabara, M., Ikeda, H., Ikebuchi, K., 2001. Serum-free coculture system for ex vivo expansion of human cord blood promitive progenitors and SCID mouse-reconstituting cells using human bone marrow primary stromal cells. Exp. Hematol., 29(2):174-182.

[45] Yin, Z.H., Liu, M., Wang, J.T., Cao, J.L., Zheng, J., 2002. Inducing rabbit bone marrow mesenchymal stem cells (MSCs) to express chondrogenic protential in vitro. Orthop. J. China, 9(6):586-589 (in Chinese).

[46] Zhang, Y., Wang, C., Liao, W., Li, Z., Guo, X., Zhao, Q., Duan, C., Xia, R., 2004a. In vitro chondrogenic phenotype differentiation of bone marrow-derived mesenchymal stem cells. J. Huazhong Univ. Sci. Technol. Med. Sci., 24(3):275-278 (in Chinese).

[47] Zhang, H., Wang, J.Z., Sun, H.Y., Zhang, J.N., Yang, S.Y., 2004b. The effects of GM1 and bFGF synergistically inducing adult rat bone marrow stromal cells to form neural progenitor cells and their differentiation. Chin. J. Traumatol., 7(1):3-6 (in Chinese).

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