Abstract
Mesenchymal stem cells (MSCs) constitute an interesting cellular source to promote brain regeneration after Parkinson’s disease. MSCs have significant advantages over other stem cell types, and greater potential for immediate clinical application. The aim of this study was to investigate whether MSCs from the human placenta could be induced to differentiate into dopaminergic cells. MSCs from the human placenta were isolated by digestion and density gradient fractionation, and their cell surface glycoproteins were analyzed by flow cytometry. These MSCs were cultured under conditions promoting differetiation into adipocytes and osteoblasts. Using a cocktail that includes basic fibroblast growth factor (bFGF), all trans retinoic acid (RA), ascorbic acid (AA) and 3-isobutyl-1-methylxanthine (IBMX), the MSCs were induced in vitro to become dopamine (DA) neurons. Then, the expression of the mRNA for the Nestin and tyrosine hydroxylase (TH) genes was assayed via RT-PCR. The expression of the Nestin, dopamine transporter (DAT), neuronal nuclear protein (NeuN) and TH proteins was determined via immunofluorescence. The synthesized and secreted DA was determined via ELISA. We found that MSCs from the human placenta exhibited a fibroblastoid morphology. Flow cytometric analyses showed that the MSCs were positive for CD44 and CD29, and negative for CD34, CD45, CD106 and HLA-DR. Moreover, they could be induced into adipocytes and osteocytes. When the MSCs were induced with bFGF, RA, AA and IBMX, they showed a change in morphology to that of neuronal-like cells. The induced cells expressed Nestin and TH mRNA, and the Nestin, DAT, NeuN and TH proteins, and synthesized and secreted DA. Our results suggest that MSCs from the human placenta have the ability to differentiate into dopaminergic cells.
Key words: Mesenchymal stem cell, Human, Placenta, Differentiation, Dopamine
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Abbreviations used
- AA
ascorbic acid
- bFGF
basic fibroblast growth factor
- DA
dopamine
- DAT
dopamine transporter
- DMEM/F12
Dulbecco’s modified Eagle’s medium/F12
- ESCs
embryonic stem cells
- FBS
fetal bovine serum
- IBMX
3-isobutyl-1-methylxanthine
- MSCs
mesenchymal stem cells
- NeuN
neuronal nuclear protein
- PBS
phosphate-buffered saline
- RA
all trans retinoic acid
- TH
tyrosine hydroxylase
References
- 1.Trzaska K.A., Kuzhikandathil E.V., Rameshwar P. Specification of a dopaminergic phenotype from adult human mesenchymal stem cells. Stem Cells. 2007;25:2797–2808. doi: 10.1634/stemcells.2007-0212. [DOI] [PubMed] [Google Scholar]
- 2.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. 2005;23:781–790. doi: 10.1634/stemcells.2004-0365. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 3.Zeng X., Cai J., Chen J., Luo Y., You Z.B., Fotter E., Wang Y., Harvey B., Miura T., Backman C., Chen G.J., Rao M.S., Freed W.J. Dopaminergic differentiation of human embryonic stem cells. Stem Cells. 2004;22:925–940. doi: 10.1634/stemcells.22-6-925. [DOI] [PubMed] [Google Scholar]
- 4.Park C.H., Minn Y.K., Lee J.Y., Choi D.H., Chang M.Y., Shim J.W., Ko J.Y., Koh H.C., Kang M.J., Kang J.S., Rhie D.J., Lee Y.S., Son H., Moon S.Y., Kim K.S., Lee S.H. In vitro and in vivo analyses of human embryonic stem cell-derived dopamine neurons. J. Neurochem. 2005;92:1265–1276. doi: 10.1111/j.1471-4159.2004.03006.x. [DOI] [PubMed] [Google Scholar]
- 5.Atmani H., Chappard D., Basle M.F. Proliferation and differentiation of osteoblasts and adipocytes in rat bone marrow stromal cell cultures: effects of dexamethasone and calcitriol. J. Cell. Biochem. 2003;89:364–372. doi: 10.1002/jcb.10507. [DOI] [PubMed] [Google Scholar]
- 6.Ai C., Todorov I., Slovak M.L., Digiusto D., Forman S.J., Shih C.C. Human marrow-derived mesodermal progenitor cells generate insulin-secreting islet-like clusters in vivo. Stem. Cells. Dev. 2007;16:757–770. doi: 10.1089/scd.2007.0038. [DOI] [PubMed] [Google Scholar]
- 7.Porter R.M., Huckle W.R., Goldstein A.S. Effect of dexamethasone withdrawal on osteoblastic differentiation of bone marrow stromal cells. J. Cell. Biochem. 2003;90:13–22. doi: 10.1002/jcb.10592. [DOI] [PubMed] [Google Scholar]
- 8.Song S., Kamath S., Mosquera D., Zigova T., Sanberg P., Vesely D.L., Sanchez-Ramos J. Expression of brain natriuretic peptide by human bone marrow stromal cells. Exp. Neurol. 2004;185:191–197. doi: 10.1016/j.expneurol.2003.09.003. [DOI] [PubMed] [Google Scholar]
- 9.Miao Z., Jin J., Chen L., Zhu J., Huang W., Zhao J., Qian H., Zhang X. Isolation of mesenchymal stem cells from human placenta: comparison with human bone marrow mesenchymal stem cells. Cell. Biol. Int. 2006;30:681–687. doi: 10.1016/j.cellbi.2006.03.009. [DOI] [PubMed] [Google Scholar]
- 10.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. 2006;24:115–124. doi: 10.1634/stemcells.2005-0053. [DOI] [PubMed] [Google Scholar]
- 11.Bibel M., Richter J., Schrenk K., Tucker K.L., Staiger V., Korte M., Goetz M., Barde Y.A. Differentiation of mouse embryonic stem cells into a defined neuronal lineage. Nat. Neurosci. 2004;7:1003–1009. doi: 10.1038/nn1301. [DOI] [PubMed] [Google Scholar]
- 12.Hsieh J., Nakashima K., Kuwabara T., Mejia E., Gage F.H. Histone deacetylase inhibition-mediated neuronal differentiation of multipotent adult neural progenitor cells. Proc. Natl. Acad. Sci. USA. 2004;101:16659–16664. doi: 10.1073/pnas.0407643101. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 13.Zhang J., Smith D., Yamamoto M., Ma L., McCaffery P. The meninges is a source of retinoic acid for the late-developing hindbrain. J. Neurosci. 2003;23:7610–7620. doi: 10.1523/JNEUROSCI.23-20-07610.2003. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 14.Lee S.H., Lumelsky N., Studer L., Auerbach J.M., McKay R.D. Efficient generation of midbrain and hindbrain neurons from mouse embryonic stem cells. Nat. Biotechnol. 2000;18:675–679. doi: 10.1038/76536. [DOI] [PubMed] [Google Scholar]
- 15.Reubinoff B.E., Itsykson P., Turetsky T., Pera M.F., Reinhartz E., Itzik A., Ben-Hur T. Neural progenitors from human embryonic stem cells. Nat. Biotechnol. 2001;19:134–140. doi: 10.1038/nbt1201-1134. [DOI] [PubMed] [Google Scholar]
- 16.Yu D.H., Lee K.H., Lee J.Y., Kim S., Shin D.M., Kim J.H., Lee Y.S., Lee Y.S., Oh S.K., Moon S.Y., Lee S.H., Lee Y.S. Changes of gene expression profiles during neuronal differentiation of central nervous system precursors treated with ascorbic acid. J. Neurosci. Res. 2004;78:29–37. doi: 10.1002/jnr.20220. [DOI] [PubMed] [Google Scholar]