Cerebrospinal fluid (CSF) contains several molecules which are essential for neurogenesis. Human dental pulp stem cells (hDPSCs) are putatively neural crest cell-derived that can differentiate into neurons and glial cells under appropriate neurotrophic factors. The aim of this study was to induce differentiation of hDPSCs into neuroglial phenotypes using retinoic acid (RA) and CSF. The hDPSCs from an impacted third molar were isolated by mechanical and digestion and cultured. The cells have treated by 10⁻⁷µM RA (RA group) for 8 days, 10% CSF (CSF group) for 8 days and RA with CSF for 8 days (RA/CSF group). Nestin, microtubule-associated protein 2 (MAP2), and glial fibrillary acidic protein immunostaining were used to examine the differentiated cells. Axonal outgrowth was detected using Bielschowsky's silver impregnation method and Nissl bodies were stained in differentiated cells by Cresyl violet. The morphology of differentiated cells in treated groups was significantly changed after 3–5 days. The results of immunocytochemistry showed the presence of neuroprogenitor marker nestin was seen in all groups. However, the high percentage of nestin positive cells and MAP2, as mature neural markers, were observed at the pre-induction and induction stage, respectively. Nissl bodies were detected as dark-blue particles in the cytoplasm of treated cells. Our findings showed the RA as pre-inducer and CSF as inducer for using in vitro differentiation of neuron-like cells and neuroglial cells from hDPSCs.
Axons ; Cerebrospinal Fluid* ; Cytoplasm ; Dental Pulp* ; Digestion ; Glial Fibrillary Acidic Protein ; Humans* ; Immunohistochemistry ; In Vitro Techniques ; Methods ; Microtubule-Associated Proteins ; Molar, Third ; Nerve Growth Factors ; Nestin ; Neural Crest ; Neurogenesis ; Neuroglia* ; Neurons ; Nissl Bodies ; Phenotype ; Silver ; Stem Cells* ; Tretinoin* ; Viola