Skip to main content
Log in

CD45CD14+CD34+ murine bone marrow low-adherent mesenchymal primitive cells preserve multilineage differentiation potential in long-term in vitro culture

  • Published:
Molecules and Cells

Abstract

Bone marrow-derived cells have been postulated as a source of multipotent mesenchymal stem cells (MSC). However, the whole fraction of MSC remains heterogeneous and the expansion of primitive subset of these cells is still not well established. Here, we optimized the protocol for propagating the low-adherent subfraction of MSC which results in long-term expansion of population characterized by CD45CD14+CD34+ phenotype along with expression of common MSC markers. We established that the expanded MSC are capable of differentiating into endothelial cells highly expressing angiogenic markers and exhibiting functional properties of endothelium. Moreover, we found these cells to be multipotent and capable of giving rise into cells from neuronal lineages. Interestingly, the expanded MSC form characteristic cellular spheres in vitro indicating primitive features of these cells. In sum, we isolated the novel multipotent subpopulation of CD45CD14+ CD34+ bone marrow-derived cells that could be maintained in long-term culture without losing this potential.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Similar content being viewed by others

References

  • Abdel-Latif, A., Bolli, R., Tleyjeh, I.M., Montori, V.M., Perin, E.C., Hornung, C.A., Zuba-Surma, E.K., Al-Mallah, M., and Dawn, B. (2007). Adult bone marrow-derived cells for cardiac repair: a systematic review and meta-analysis. Arch. Intern. Med. 167, 989–997.

    Article  PubMed  Google Scholar 

  • Alviano, F., Fossati, V., Marchionni, C., Arpinati, M., Bonsi, L., Franchina, M., Lanzoni, G., Cantoni, S., Cavallini, C., Bianchi, F., et al. (2007). Term Amniotic membrane is a high throughput source for multipotent Mesenchymal Stem Cells with the ability to differentiate into endothelial cells in vitro. BMC Dev. Biol. 7, 11.

    Article  PubMed  Google Scholar 

  • Beyer Nardi, N., and da Silva Meirelles, L., (2006). Mesenchymal stem cells: isolation, in vitro expansion and characterization. Handb. Exp. Pharmacol. 174, 249–282.

    PubMed  Google Scholar 

  • Brunt, K.R., Hall, S.R.R., Ward, C.A., and Melo, L.G. (2007). Endothelial progenitor cell and mesenchymal stem cell isolation, characterization, viral transduction. Methods Mol. Med. 139, 197–210.

    Article  PubMed  CAS  Google Scholar 

  • Chawla, A., Schwarz, E.J., Dimaculangan, D.D., and Lazar, M.A. (1994). Peroxisome proliferator-activated receptor (PPAR) gamma: adipose-predominant expression and induction early in adipocyte differentiation. Endocrinology 135, 798–800.

    Article  PubMed  CAS  Google Scholar 

  • Chen, M., Lie, P., Li, Z., and Wei, X. (2009). Endothelial differentiation of Wharton’s jelly-derived mesenchymal stem cells in comparison with bone marrow-derived mesenchymal stem cells. Exp. Hematol. 37, 629–640.

    Article  PubMed  CAS  Google Scholar 

  • Cleaver, O., and Melton, D.A. (2003). Endothelial signaling during development. Nat. Med. 9, 661–668.

    Article  PubMed  CAS  Google Scholar 

  • Copland, I., Sharma, K., Lejeune, L., Eliopoulos, N., Stewart, D., Liu, P., Lachapelle, K., and Galipeau, J. (2008). CD34 expression on murine marrow-derived mesenchymal stromal cells: impact on neovascularization. Exp. Hematol. 36, 93–103.

    Article  PubMed  CAS  Google Scholar 

  • Dawn, B., Tiwari, S., Kucia, M.J., Zuba-Surma, E.K., Guo, Y., Sanganalmath, S.K., Abdel-Latif, A., Hunt, G., Vincent, R.J., Taher, H., et al. (2008). Transplantation of bone marrow-derived very small embryonic-like stem cells attenuates left ventricular dysfunction and remodeling after myocardial infarction. Stem Cells 26, 1646–1655.

    Article  PubMed  Google Scholar 

  • Dawn, B., Abdel-Latif, A., Sanganalmath, S.K., Flaherty, M.P., Zuba-Surma, E.K. (2009). Cardiac repair with adult bone marrowderived cells: the clinical evidence. Antioxid. Redox Signal. 11, 1865–1882.

    Article  PubMed  CAS  Google Scholar 

  • D’Ippolito, G., Diabira, S., Howard, G.A., Menei, P., Roos, B.A., and Schiller, P.C. (2004). Marrow-isolated adult multilineage inducible (MIAMI) cells, a unique population of postnatal young and old human cells with extensive expansion and differentiation potential. J. Cell Sci. 117, 2971–2981.

    Article  PubMed  Google Scholar 

  • Fei, R.G., Penn, P.E., and Wolf, N.S. (1990). A method to establish pure fibroblast and endothelial cell colony cultures from murine bone marrow. Exp. Hematol. 18, 953–957.

    PubMed  CAS  Google Scholar 

  • Fernandez Pujol, B., Lucibello, F.C., Gehling, U.M., Lindemann, K., Weidner, N., Zuzarte, M.L., Adamkiewicz, J., Elsässer, H.P., Müller, R., and Havemann, K. (2000). Endothelial-like cells derived from human CD14 positive monocytes. Differentiation 65, 287–300.

    Article  PubMed  CAS  Google Scholar 

  • Gang, E.J., Jeong, J.A., Han, S., Yan, Q., Jeon, C., and Kim, H. (2006). In vitro endothelial potential of human UC blood-derived mesenchymal stem cells. Cytotherapy 8, 215–227.

    Article  PubMed  CAS  Google Scholar 

  • Girdlestone, J., Limbani, V., Cutler, A., and Navarrete, C. (2009). Efficient expansion of mesenchymal stromal cells from umbilical cord under low serum conditions. Cytotherapy [Epub ahead of print].

  • Gnecchi, M., and Melo, L.G. (2009). Bone marrow-derived mesenchymal stem cells: isolation, expansion, characterization, viral transduction, and production of conditioned medium. Methods Mol. Biol. 482, 281–294.

    Article  PubMed  CAS  Google Scholar 

  • Grove, J.E., Bruscia, E., and Krause, D.S. (2004). Plasticity of bone marrow-derived stem cells. Stem Cells 22, 487–500.

    Article  PubMed  Google Scholar 

  • Hofstetter, C.P., Schwarz, E.J., Hess, D., Widenfalk, J., El Manira, A., Prockop, D.J., and Olson, L. (2002). Marrow stromal cells form guiding strands in the injured spinal cord and promote recovery. Proc. Natl. Acad. Sci. USA 99, 2199–2204.

    Article  PubMed  CAS  Google Scholar 

  • Houlihan, D.D., and Newsome, P.N. (2008). Critical review of clinical trials of bone marrow stem cells in liver disease. Gastroenterology 135, 438–450.

    Article  PubMed  CAS  Google Scholar 

  • Ingram, D.A., Mead, L.E., Tanaka, H., Meade, V., Fenoglio, A., Mortell, K., Pollok, K., Ferkowicz, M.J., Gilley, D., and Yoder, M.C. (2004). Identification of a novel hierarchy of endothelial progenitor cells using human peripheral and umbilical cord blood. Blood 104, 2752–2760.

    Article  PubMed  CAS  Google Scholar 

  • Jain, R.K. (2003). Molecular regulation of vessel maturation. Nat. Med. 9, 685–693.

    Article  PubMed  CAS  Google Scholar 

  • 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, 41–49.

    Article  PubMed  CAS  Google Scholar 

  • Kabos, P., Ehtesham, M., Kabosova, A., Black, K.L., and Yu, J.S. (2002). Generation of neural progenitor cells from whole adult bone marrow. Exp. Neurol. 178, 288–293.

    Article  PubMed  CAS  Google Scholar 

  • Kaiser, S., Hackanson, B., Follo, M., Mehlhorn, A., Geiger, K., Ihorst, G., and Kapp, U. (2007). BM cells giving rise to MSC in culture have a heterogeneous CD34 and CD45 phenotype. Cytotherapy 9, 439–450.

    Article  PubMed  CAS  Google Scholar 

  • Karussis, D., Kassis, I., Kurkalli, B.G.S., and Slavin, S. (2008). Immunomodulation and neuroprotection with mesenchymal bone marrow stem cells (MSCs): a proposed treatment for multiple sclerosis and other neuroimmunological/neurodegenerative diseases. J. Neurol. Sci. 265, 131–135.

    Article  PubMed  CAS  Google Scholar 

  • Kopher, R.A., Penchev, V.R., Islam, M.S., Hill, K.L., Khosla, S., and Kaufman, D.S. (2010). Human embryonic stem cell-derived CD34+ cells function as MSC progenitor cells. Bone 47, 718–728.

    Article  PubMed  CAS  Google Scholar 

  • Kucia, M., Reca, R., Campbell, F.R., Zuba-Surma, E., Majka, M., Ratajczak, J., and Ratajczak, M.Z. (2006). A population of very small embryonic-like (VSEL) CXCR4(+)SSEA-1(+)Oct-4+ stem cells identified in adult bone marrow. Leukemia 20, 857–869.

    Article  PubMed  CAS  Google Scholar 

  • Kucia, M., Wysoczynski, M., Ratajczak, J., and Ratajczak, M.Z. (2008). Identification of very small embryonic like (VSEL) stem cells in bone marrow. Cell Tissue Res. 331, 125–134.

    Article  PubMed  CAS  Google Scholar 

  • Kuwana, M., Okazaki, Y., Kodama, H., Izumi, K., Yasuoka, H., Ogawa, Y., Kawakami, Y., and Ikeda, Y. (2003). Human circulating CD14+ monocytes as a source of progenitors that exhibit mesenchymal cell differentiation. J. Leukoc. Biol. 74, 833–845.

    Article  PubMed  CAS  Google Scholar 

  • Li, Y., Zhang, C., Xiong, F., Yu, M., Peng, F., Shang, Y., Zhao, C., Xu, Y., Liu, Z., Zhou, C., et al. (2008). Comparative study of mesenchymal stem cells from C57BL/10 and mdx mice. BMC Cell Biol. 9, 24.

    Article  PubMed  Google Scholar 

  • Lim, S.Y., Kim, Y.S., Ahn, Y., Jeong, M.H., Hong, M.H., Joo, S.Y., Nam, K.I., Cho, J.G., Kang, P.M., and Park, J.C. (2006). The effects of mesenchymal stem cells transduced with Akt in a porcine myocardial infarction model. Cardiovasc. Res. 70, 530–542.

    Article  PubMed  CAS  Google Scholar 

  • Martin-Rendon, E., Brunskill, S.J., Hyde, C.J., Stanworth, S.J., Mathur, A., and Watt, S.M. (2008). Autologous bone marrow stem cells to treat acute myocardial infarction: a systematic review. Eur. Heart J. 29, 1807–1818.

    Article  PubMed  CAS  Google Scholar 

  • Matoba, S., Tatsumi, T., Murohara, T., Imaizumi, T., Katsuda, Y., Ito, M., Saito, Y., Uemura, S., Suzuki, H., Fukumoto, S., et al. (2008). Long-term clinical outcome after intramuscular implantation of bone marrow mononuclear cells (therapeutic angiogenesis by cell transplantation [TACT] trial) in patients with chronic limb ischemia. Am. Heart J. 156, 1010–1018.

    Article  PubMed  Google Scholar 

  • Oswald, J., Boxberger, S., Jørgensen, B., Feldmann, S., Ehninger, G., Bornhäuser, M., and Werner, C. (2004). Mesenchymal stem cells can be differentiated into endothelial cells in vitro. Stem Cells 22, 377–384.

    Article  PubMed  Google Scholar 

  • Phinney, D.G., and Prockop, D.J. (2007). Concise review: mesenchymal stem/multipotent stromal cells: the state of transdifferentiation and modes of tissue repair-current views. Stem Cells 25, 2896–2902.

    Article  PubMed  Google Scholar 

  • Pitas, R.E., Boyles, J., Mahley, R.W., and Bissell, D.M. (1985). Uptake of chemically modified low density lipoproteins in vivo is mediated by specific endothelial cells. J. Cell Biol. 100, 103–117.

    Article  PubMed  CAS  Google Scholar 

  • 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., and Marshak, D.R. (1999). Multilineage potential of adult human mesenchymal stem cells. Science 284, 143–147.

    Article  PubMed  CAS  Google Scholar 

  • Quirici, N., Soligo, D., Caneva, L., Servida, F., Bossolasco, P., and Deliliers, G.L. (2001). Differentiation and expansion of endothelial cells from human bone marrow CD133(+) cells. Br. J. Haematol. 115, 186–94.

    Article  PubMed  CAS  Google Scholar 

  • Reynolds, B.A., and Weiss, S. (1992). Generation of neurons and astrocytes from isolated cells of the adult mammalian central nervous system. Science 255, 1707–1710.

    Article  PubMed  CAS  Google Scholar 

  • Romagnani, P., Annunziato, F., Liotta, F., Lazzeri, E., Mazzinghi, B., Frosali, F., Cosmi, L., Maggi, L., Lasagni, L., Scheffold, A., et al. (2005). CD14+CD34 low cells with stem cell phenotypic and functional features are the major source of circulating endothelial progenitors. Circ. Res. 97, 314–322.

    Article  PubMed  CAS  Google Scholar 

  • Shiota, M., Heike, T., Haruyama, M., Baba, S., Tsuchiya, A., Fujino, H., Kobayashi, H., Kato, T., Umeda, K., Yoshimoto, M., et al. (2007). Isolation and characterization of bone marrow-derived mesenchymal progenitor cells with myogenic and neuronal properties. Exp. Cell Res. 313, 1008–1023.

    Article  PubMed  CAS  Google Scholar 

  • Torrente, Y., and Polli, E. (2008). Mesenchymal stem cell transplantation for neurodegenerative diseases. Cell Transplant 17, 1103–1113.

    Article  PubMed  Google Scholar 

  • Wang, Q.R., Wang, B.H., Huang, Y.H., Dai, G., Li, W.M., and Yan, Q. (2008). Purification and growth of endothelial progenitor cells from murine bone marrow mononuclear cells. J. Cell Biochem. 103, 21–29.

    Article  PubMed  CAS  Google Scholar 

  • Zhang, S.J., Zhang, H., Hou, M., Zheng, Z., Zhou, J., Su, W., Wei, Y., and Hu, S. (2007). Is it possible to obtain “true endothelial progenitor cells” by in vitro culture of bone marrow mononuclear cells? Stem Cells Dev. 16, 683–690.

    Article  PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Jozef Dulak.

About this article

Cite this article

Szade, K., Zuba-Surma, E., Rutkowski, A.J. et al. CD45CD14+CD34+ murine bone marrow low-adherent mesenchymal primitive cells preserve multilineage differentiation potential in long-term in vitro culture. Mol Cells 31, 497–507 (2011). https://doi.org/10.1007/s10059-011-2176-y

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10059-011-2176-y

Keywords

Navigation