Hindawi Publishing Corporation Stem Cells International Volume 2013, Article ID 250740, 10 pages http://dx.doi.org/10.1155/2013/250740 Research Article Differentiation of Dental Pulp Stem Cells into Neuron-Like Cells in Serum-Free Medium Shahrul Hisham Zainal Ariffin, 1 Shabnam Kermani, 1 Intan Zarina Zainol Abidin, 1 Rohaya Megat Abdul Wahab, 2 Zulham Yamamoto, 1 Sahidan Senafi, 1 Zaidah Zainal Ariffin, 3 and Mohamad Abdul Razak 4 1 School of Biosciences and Biotechnology, Faculty of Science and Technology, Universiti Kebangsaan Malaysia, 43600 Bangi, Selangor, Malaysia 2 Department of Orthodontics, Faculty of Dentistry, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur, Malaysia 3 School of Biology, Faculty of Applied Science, Universiti Teknologi MARA, 40450 Shah Alam, Selangor, Malaysia 4 Allianze University College of Medical Sciences, Waziria Medical Square, Jalan Bertam 2, 13200 Kepala Batas, Penang, Malaysia Correspondence should be addressed to Shahrul Hisham Zainal Arifn; shahroy8@gmail.com Received 13 December 2012; Revised 23 September 2013; Accepted 7 October 2013 Academic Editor: Deepa Bhartiya Copyright © 2013 Shahrul Hisham Zainal Arifn et al. Tis is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Dental pulp tissue contains dental pulp stem cells (DPSCs). Dental pulp cells (also known as dental pulp-derived mesenchymal stem cells) are capable of diferentiating into multilineage cells including neuron-like cells. Te aim of this study was to examine the capability of DPSCs to diferentiate into neuron-like cells without using any reagents or growth factors. DPSCs were isolated from teeth extracted from 6- to 8-week-old mice and maintained in complete medium. Te cells from the fourth passage were induced to diferentiate by culturing in medium without serum or growth factors. RT-PCR molecular analysis showed characteristics of Cd146 + , Cd166 + , and Cd31 − in DPSCs, indicating that these cells are mesenchymal stem cells rather than hematopoietic stem cells. Afer 5 days of neuronal diferentiation, the cells showed neuron-like morphological changes and expressed MAP2 protein. Te activation of Nestin was observed at low level prior to diferentiation and increased afer 5 days of culture in diferentiation medium, whereas Tub3 was activated only afer 5 days of neuronal diferentiation. Te proliferation of the diferentiated cells decreased in comparison to that of the control cells. Dental pulp stem cells are induced to diferentiate into neuron-like cells when cultured in serum- and growth factor-free medium. 1. Introduction Dental pulp tissue contains many types of cells including committed cells (e.g., endothelial cells) and uncommitted cells (i.e., DPSCs). DPSCs are of mesenchymal stem cells (MSCs) [1]. In mice, the majority of MSCs were isolated from bone marrow [2] and peripheral blood [3, 4]. Tese MSCs can be characterized by the expression of specifc gene markers such as CD44, CD73, CD90, CD105, CD117, and CD166 [5, 6]. DPSCs are capable of diferentiating into multilineage cells [7–9] including neuron-like cells [10]. Neuron-like cells diferentiated from MSCs derived from bone marrow cells [11–13] and brain [14]. However, MSCs derived from dental pulp, that is, DPSCs, are also capable of diferentiating into neuron-like cells [10]. Te characteristics of MSCs from bone marrow are similar to those cells derived from dental pulp [11]. Both types of MSCs express Cd44, Cd106, Cd146, and Cd166 [15–17]. Many factors are involved in neuronal diferentiation including nestin [18], tubulin3 (Tub3) [19], and MAP2 [20]. Nestin is involved in the radial growth of axons during neuronal diferentiation in vertebrate cells [19, 21]. Terefore, Nestin is known as a neural marker and its presence can be considered as a criterion for the ability to diferentiate