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Directed differentiation of aged human bone marrow multipotent stem cells effectively generates dopamine neurons

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Abstract

This study aimed to isolate aged human bone marrow multipotent stem cells (hAMSCs) with the potential for multilineage differentiation and to directly induce the cells to generate dopamine neurons, which could be used for Parkinson's disease therapy. We compared different culture methods for stem cells from aged human bone marrow and identified hAMSCs that could proliferate in vitro for at least 60 doubling times. Using RT-PCR and IHC, we found that these hAMSCs expressed pluripotent genes, such as Oct4, Sox2, and Nanog. In vitro studies also proved that hAMSCs could differentiate into three germ layer-derived cell types, such as osteogenic, chondrogenic, adipogenic, and hepatocyte-liked cells. After induction for more than 20 d in vitro with retinoic acid, basic fibroblast growth factor, and sonic hedgehog using a two-step method and withdrawal of serum, hAMSCs could differentiate into dopamine neurons at the positive ratio of 70%, which showed DA secretion function upon depolarization. In conclusion, we suggest that hAMSCs can be used as cell sources to develop medical treatments to prevent the progression of Parkinson's disease, especially in aged persons.

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References

  • Baquet Z. C.; Bickford P. C.; Jones K. R. Brain-derived neurotrophic factor is required for the establishment of the proper number of dopaminergic neurons in the substantia nigra pars compacta. J. Neurosci. 25: 6251–9; 2005.

    Article  PubMed  CAS  Google Scholar 

  • Barry F.; Boynton R.; Murphy M.; Haynesworth S.; Zaia J. The SH-3 and SH-4 antibodies recognize distinct epitopes on CD73 from human mesenchymal stem cells. Biochem. Biophys. Res. Commun. 289: 519–24; 2001.

    Article  PubMed  CAS  Google Scholar 

  • Betarbet R.; Sherer T. B.; Greenamyre J. T. Animal models of Parkinson's disease. Bioessays 24: 308–18; 2002.

    Article  PubMed  CAS  Google Scholar 

  • Boyer L. A.; Lee T. I.; Cole M. F.; Johnstone S. E.; Levine S. S.; Zucker J. P.; Guenther M. G.; Kumar R. M.; Murray H. L.; Jenner R. G. et al. Core transcriptional regulatory circuitry in human embryonic stem cells. Cell 122: 947–56; 2005.

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Buytaert-Hoefen K. A.; Alvarez E.; Freed C. R. Generation of tyrosine hydroxylase positive neurons from human embryonic stem cells after coculture with cellular substrates and exposure to GDNF. Stem Cells 22: 669–74; 2004.

    Article  PubMed  CAS  Google Scholar 

  • Cauffman G.; Liebaers I.; Van Steirteghem A.; Van de Velde H. POU5F1 isoforms show different expression patterns in human embryonic stem cells and preimplantation embryos. Stem Cells 24: 2685–91; 2006.

    Article  PubMed  CAS  Google Scholar 

  • Colter D. C.; Class R.; DiGirolamo C. M.; Prockop D. J. Rapid expansion of recycling stem cells in cultures of plastic-adherent cells from human bone marrow. Proc. Natl. Acad. Sci. U. S. A. 97: 3213–8; 2000.

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Cooper O.; Hargus G.; Deleidi M.; Blak A.; Osborn T.; Marlow E.; Lee K.; Levy A.; Perez-Torres E.; Yow A. et al. Differentiation of human ES and Parkinson's disease iPS cells into ventral midbrain dopaminergic neurons requires a high activity form of SHH, FGF8a and specific regionalization by retinoic acid. Mol. Cell. Neurosci. 45: 258–66; 2010.

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • D'Ippolito G.; Diabira S.; Howard G. A.; Menei P.; Roos B. A.; Schiller P. C. 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: 2971–81; 2004.

    Article  PubMed  CAS  Google Scholar 

  • Dezawa M.; Kanno H.; Hoshino M.; Cho H.; Matsumoto N.; Itokazu Y.; Tajima N.; Yamada H.; Sawada H.; Ishikawa H. et al. Specific induction of neuronal cells from bone marrow stromal cells and application for autologous transplantation. J. Clin. Invest. 113: 1701–10; 2004.

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Emborg M. E.; Zhang Z., Joers V.; Brunner K.; Bondarenko V.; Ohshima S.; Zhang S. C. Intracerebral Transplantation of differentiated human embryonic stem cells to hemiparkinsonian monkeys. Cell Transplant (in press); 2013.

  • Erceg S.; Ronaghi M.; Stojkovic M. Human embryonic stem cell differentiation toward regional specific neural precursors. Stem Cells 27: 78–87; 2009.

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Fu Y. S.; Cheng Y. C.; Lin M. Y.; Cheng H.; Chu P. M.; Chou S. C.; Shih Y. H.; Ko M. H.; Sung M. S. Conversion of human umbilical cord mesenchymal stem cells in Wharton's jelly to dopaminergic neurons in vitro: potential therapeutic application for Parkinsonism. Stem Cells 24: 115–24; 2006.

    Article  PubMed  Google Scholar 

  • Fuccillo M.; Joyner A. L.; Fishell G. Morphogen to mitogen: the multiple roles of hedgehog signalling in vertebrate neural development. Nat. Rev. Neurosci. 7: 772–83; 2006.

    Article  PubMed  CAS  Google Scholar 

  • Granholm A. C.; Reyland M.; Albeck D.; Sanders L.; Gerhardt G.; Hoernig G.; Shen L.; Westphal H.; Hoffer B. Glial cell line-derived neurotrophic factor is essential for postnatal survival of midbrain dopamine neurons. J. Neurosci. 20: 3182–90; 2000.

    PubMed  CAS  Google Scholar 

  • Hargus G.; Cooper O.; Deleidi M.; Levy A.; Lee K.; Marlow E.; Yow A.; Soldner F.; Hockemeyer D.; Hallett P. J. et al. Differentiated Parkinson patient-derived induced pluripotent stem cells grow in the adult rodent brain and reduce motor asymmetry in Parkinsonian rats. Proc. Natl. Acad. Sci. U. S. A. 107: 15921–6; 2010.

    Article  PubMed Central  PubMed  Google Scholar 

  • Hayashi T.; Wakao S.; Kitada M.; Ose T.; Watabe H.; Kuroda Y.; Mitsunaga K.; Matsuse D.; Shigemoto T.; Ito A. et al. Autologous mesenchymal stem cell-derived dopaminergic neurons function in parkinsonian macaques. J. Clin. Invest. 123: 272–84; 2012.

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Ingham P. W.; Placzek M. Orchestrating ontogenesis: variations on a theme by sonic hedgehog. Nat. Rev. Genet. 7: 841–50; 2006.

    Article  PubMed  CAS  Google Scholar 

  • Jiang Y.; Henderson D.; Blackstad M.; Chen A.; Miller R. F.; Verfaillie C. M. Neuroectodermal differentiation from mouse multipotent adult progenitor cells. Proc. Natl. Acad. Sci. U. S. A. 100(Suppl 1): 11854–60; 2003.

    Article  PubMed Central  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. Pluripotency of mesenchymal stem cells derived from adult marrow. Nature 418: 41–9; 2002.

    Article  PubMed  CAS  Google Scholar 

  • Korecka J. A.; Verhaagen J.; Hol E. M. Cell-replacement and gene-therapy strategies for Parkinson's and Alzheimer's disease. Regen. Med. 2: 425–46; 2007.

    Article  PubMed  CAS  Google Scholar 

  • Lee S.; Lee B.; Lee J. W.; Lee S. K. Retinoid signaling and neurogenin2 function are coupled for the specification of spinal motor neurons through a chromatin modifier CBP. Neuron 62: 641–54; 2009.

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Lewitzky M.; Yamanaka S. Reprogramming somatic cells towards pluripotency by defined factors. Curr. Opin. Biotechnol. 18: 467–73; 2007.

    Article  PubMed  CAS  Google Scholar 

  • Li J. Y.; Christophersen N. S.; Hall V.; Soulet D.; Brundin P. Critical issues of clinical human embryonic stem cell therapy for brain repair. Trends Neurosci. 31: 146–53; 2008.

    Article  PubMed  CAS  Google Scholar 

  • Marti E.; Bovolenta P. Sonic hedgehog in CNS development: one signal, multiple outputs. Trends Neurosci. 25: 89–96; 2002.

    Article  PubMed  CAS  Google Scholar 

  • Mavromatakis Y. E.; Lin W.; Metzakopian E.; Ferri A. L.; Yan C. H.; Sasaki H.; Whisett J.; Ang S. L. Foxa1 and Foxa2 positively and negatively regulate Shh signalling to specify ventral midbrain progenitor identity. Mech. Dev. 128: 90–103; 2010.

    Article  PubMed  CAS  Google Scholar 

  • Murase S.; McKay R. D. A specific survival response in dopamine neurons at most risk in Parkinson's disease. J. Neurosci. 26: 9750–60; 2006.

    Article  PubMed  CAS  Google Scholar 

  • Nauta A. J.; Fibbe W. E. Immunomodulatory properties of mesenchymal stromal cells. Blood 110: 3499–506; 2007.

    Article  PubMed  CAS  Google Scholar 

  • Perrier A. L.; Tabar V.; Barberi T.; Rubio M. E.; Bruses J.; Topf N.; Harrison N. L.; Studer L. Derivation of midbrain dopamine neurons from human embryonic stem cells. Proc. Natl. Acad. Sci. U. S. A. 101: 12543–8; 2004.

    Article  PubMed Central  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.; Marshak D. R. Multilineage potential of adult human mesenchymal stem cells. Science 284: 143–7; 1999.

    Article  PubMed  CAS  Google Scholar 

  • Reyes M.; Lund T.; Lenvik T.; Aguiar D.; Koodie L.; Verfaillie C. M. Purification and ex vivo expansion of postnatal human marrow mesodermal progenitor cells. Blood 98: 2615–25; 2001.

    Article  PubMed  CAS  Google Scholar 

  • Roy N. S.; Cleren C.; Singh S. K.; Yang L.; Beal M. F.; Goldman S. A. Functional engraftment of human ES cell-derived dopaminergic neurons enriched by coculture with telomerase-immortalized midbrain astrocytes. Nat. Med. 12: 1259–68; 2006.

    Article  PubMed  CAS  Google Scholar 

  • Smidt M. P.; Burbach J. P. How to make a mesodiencephalic dopaminergic neuron. Nat. Rev. Neurosci. 8: 21–32; 2007.

    Article  PubMed  CAS  Google Scholar 

  • Snyder B. J.; Olanow C. W. Stem cell treatment for Parkinson's disease: an update for 2005. Curr. Opin. Neurol. 18: 376–85; 2005.

    Article  PubMed  CAS  Google Scholar 

  • Suon S.; Yang M.; Iacovitti L. Adult human bone marrow stromal spheres express neuronal traits in vitro and in a rat model of Parkinson's disease. Brain Res. 1106: 46–51; 2006.

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Tatard V. M.; D'Ippolito G.; Diabira S.; Valeyev A.; Hackman J.; McCarthy M.; Bouckenooghe T.; Menei P.; Montero-Menei C. N.; Schiller P. C. Neurotrophin-directed differentiation of human adult marrow stromal cells to dopaminergic-like neurons. Bone 40: 360–73; 2007.

    Article  PubMed  CAS  Google Scholar 

  • Tremain N.; Korkko J.; Ibberson D.; Kopen G. C.; DiGirolamo C.; Phinney D. G. MicroSAGE analysis of 2,353 expressed genes in a single cell-derived colony of undifferentiated human mesenchymal stem cells reveals mRNAs of multiple cell lineages. Stem Cells 19: 408–18; 2001.

    Article  PubMed  CAS  Google Scholar 

  • Trzaska K. A.; Kuzhikandathil E. V.; Rameshwar P. Specification of a dopaminergic phenotype from adult human mesenchymal stem cells. Stem Cells 25: 2797–808; 2007.

    Article  PubMed  CAS  Google Scholar 

  • Trzaska K. A.; Rameshwar P. Current advances in the treatment of Parkinson's disease with stem cells. Curr. Neurovasc. Res. 4: 99–109; 2007.

    Article  PubMed  CAS  Google Scholar 

  • Ulloa-Montoya F.; Verfaillie C. M.; Hu W. S. Culture systems for pluripotent stem cells. J. Biosci. Bioeng. 100: 12–27; 2005.

    Article  PubMed  CAS  Google Scholar 

  • Wakeman D. R.; Weiss S.; Sladek J. R.; Elsworth J. D.; Bauereis B.; Leranth C.; Hurley P.J.; Roth R.H.; Redmond E. Survival and integration of neurons derived from human embryonic stem cells in MPTP lesioned primates. Cell Transplant (2013); 2013.

  • Wernig M.; Zhao J. P.; Pruszak J.; Hedlund E.; Fu D.; Soldner F.; Broccoli V.; Constantine-Paton M.; Isacson O.; Jaenisch R. Neurons derived from reprogrammed fibroblasts functionally integrate into the fetal brain and improve symptoms of rats with Parkinson's disease. Proc. Natl. Acad. Sci. U. S. A. 105: 5856–61; 2008.

    Article  PubMed Central  PubMed  Google Scholar 

  • Wu M.; Zhang Y.; Wu N. H.; Shen Y. F. Histone marks and chromatin remodelers on the regulation of neurogenin1 gene in RA induced neuronal differentiation of P19 cells. J. Cell. Biochem. 107: 264–71; 2009.

    Article  PubMed  CAS  Google Scholar 

  • Yan Y.; Yang D.; Zarnowska E. D.; Du Z.; Werbel B.; Valliere C.; Pearce R. A.; Thomson J. A.; Zhang S. C. Directed differentiation of dopaminergic neuronal subtypes from human embryonic stem cells. Stem Cells 23: 781–90; 2005.

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Yoon Y. S.; Wecker A.; Heyd L.; Park J. S.; Tkebuchava T.; Kusano K.; Hanley A.; Scadova H.; Qin G.; Cha D. H. et al. Clonally expanded novel multipotent stem cells from human bone marrow regenerate myocardium after myocardial infarction. J. Clin. Invest. 115: 326–38; 2005.

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  • Zhang Z. X.; Roman G. C.; Hong Z.; Wu C. B.; Qu Q. M.; Huang J. B.; Zhou B.; Geng Z. P.; Wu J. X.; Wen H. B. et al. Parkinson's disease in China: prevalence in Beijing, Xian, and Shanghai. Lancet 365: 595–7; 2005.

    Article  PubMed  Google Scholar 

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Acknowledgments

This work was supported by National 973 Project of China (no. 2011CB965101) and Major Program of The Eleventh Five-Year Plan of PLA Medicine (no. 06G62)

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Authors and Affiliations

  1. Research Center of Developmental Biology, Second Military Medical University, XiangYin road 800, Shanghai, China

    Lixing Fan, Kaimeng Hu, Kaihong Ji, Qing Sun, Jun Xiong, Ling Yang & Houqi Liu

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  1. Lixing Fan
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  2. Kaimeng Hu
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  4. Qing Sun
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Correspondence to Houqi Liu.

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Editor: T. Okamoto

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Fan, L., Hu, K., Ji, K. et al. Directed differentiation of aged human bone marrow multipotent stem cells effectively generates dopamine neurons. In Vitro Cell.Dev.Biol.-Animal 50, 304–312 (2014). https://doi.org/10.1007/s11626-013-9701-6

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