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J Korean Med Sci. 2016 Feb;31(2):171-7. doi: 10.3346/jkms.2016.31.2.171. Epub 2016 Jan 13.

Differentiation of Human Dental Pulp Stem Cells into Dopaminergic Neuron-like Cells in Vitro.

Author information

1
BioMedical Research Institute, Kyungpook National University Hospital, Daegu, Korea.
2
Wake Forest Institute for Regenerative Medicine, Wake Forest University School of Medicine, Winston-Salem, NC, USA.
3
Department of Neural Development and Disease, Korea Brain Research Institute, Daegu, Korea.
4
Department of Urology, Kyungpook National University, Daegu, Korea.

Abstract

We investigated the potential of human dental pulp stem cells (hDPSCs) to differentiate into dopaminergic neurons in vitro as an autologous stem cell source for Parkinson's disease treatment. The hDPSCs were expanded in knockout-embryonic stem cell (KO-ES) medium containing leukemia inhibitory factor (LIF) on gelatin-coated plates for 3-4 days. Then, the medium was replaced with KO-ES medium without LIF to allow the formation of the neurosphere for 4 days. The neurosphere was transferred into ITS medium, containing ITS (human insulin-transferrin-sodium) and fibronectin, to select for Nestin-positive cells for 6-8 days. The cells were then cultured in N-2 medium containing basic fibroblast growth factor (FGF), FGF-8b, sonic hedgehog-N, and ascorbic acid on poly-l-ornithine/fibronectin-coated plates to expand the Nestin-positive cells for up to 2 weeks. Finally, the cells were transferred into N-2/ascorbic acid medium to allow for their differentiation into dopaminergic neurons for 10-15 days. The differentiation stages were confirmed by morphological, immunocytochemical, flow cytometric, real-time PCR, and ELISA analyses. The expressions of mesenchymal stem cell markers were observed at the early stages. The expressions of early neuronal markers were maintained throughout the differentiation stages. The mature neural markers showed increased expression from stage 3 onwards. The percentage of cells positive for tyrosine hydroxylase was 14.49%, and the amount was 0.526 ± 0.033 ng/mL at the last stage. hDPSCs can differentiate into dopaminergic neural cells under experimental cell differentiation conditions, showing potential as an autologous cell source for the treatment of Parkinson's disease.

KEYWORDS:

Autologous Stem Cell Therapy; Dopaminergic Differentiation; Dopaminergic Neural Cells; Human Dental Pulp Stem Cells; Parkinson Disease; Tyrosine Hydroxylase

PMID:
26839468
PMCID:
PMC4729494
DOI:
10.3346/jkms.2016.31.2.171
[Indexed for MEDLINE]
Free PMC Article

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