title: Neurogenic maturation of human dental pulp stem cells following neurosphere generation induces morphological and electrophysiological characteristics of functional neurons.
abstract: Cell-based therapies are emerging as an alternative treatment option to promote functional recovery in patients suffering from neurological disorders, which are the major cause of death and permanent disability. The present study aimed to differentiate human dental pulp stem cells (hDPSCs) toward functionally active neuronal cells in vitro. hDPSCs were subjected to a two-step protocol. First, neuronal induction was acquired through the formation of neurospheres, followed by neuronal maturation, based on cAMP and neurotrophin-3 (NT-3) signaling. At the ultrastructural level, it was shown that the intra-spheral microenvironment promoted intercellular communication. hDPSCs grew out of the neurospheres in vitro and established a neurogenic differentiated hDPSC culture (d-hDPSCs) upon cAMP and NT-3 signaling. d-hDPSCs were characterized by the increased expression of neuronal markers such as neuronal nuclei, microtubule-associated protein 2, neural cell adhesion molecule, growth-associated protein 43, synapsin I, and synaptophysin compared with nondifferentiated hDPSCs. Enzyme-linked immunosorbent assay demonstrated that the secretion of brain-derived neurotrophic factor, vascular endothelial growth factor, and nerve growth factor differed between d-hDPSCs and hDPSCs. d-hDPSCs acquired neuronal features, including multiple intercommunicating cytoplasmic extensions and increased vesicular transport, as shown by the electron microscopic observation. Patch clamp analysis demonstrated the functional activity of d-hDPSCs by the presence of tetrodotoxin- and tetraethyl ammonium-sensitive voltage-gated sodium and potassium channels, respectively. A subset of d-hDPSCs was able to fire a single action potential. The results reported in this study demonstrate that hDPSCs are capable of neuronal commitment following neurosphere formation, characterized by distinct morphological and electrophysiological properties of functional neuronal cells.
authors.affiliation: 1 Group of Morphology, Biomedical Research Institute, Hasselt University , Diepenbeek, Belgium .
authors.forename: Pascal
authors.initials: P
authors.lastname: Gervois
authors.forename: Tom
authors.initials: T
authors.lastname: Struys
authors.forename: Petra
authors.initials: P
authors.lastname: Hilkens
authors.forename: Annelies
authors.initials: A
authors.lastname: Bronckaers
authors.forename: Jessica
authors.initials: J
authors.lastname: Ratajczak
authors.forename: Constantinus
authors.initials: C
authors.lastname: Politis
authors.forename: Bert
authors.initials: B
authors.lastname: Brône
authors.forename: Ivo
authors.initials: I
authors.lastname: Lambrichts
authors.forename: Wendy
authors.initials: W
authors.lastname: Martens
mesh_terms: D000200:Action Potentials; D000293:Adolescent; D000328:Adult; D002450:Cell Communication; D018929:Cell Culture Techniques; D002469:Cell Separation / Q000379:methods; D048430:Cell Shape; D002478:Cells, Cultured; D060833:Cellular Microenvironment; D003470:Culture Media / Q000494:pharmacology; D003782:Dental Pulp / Q000166:cytology*; D005260:Female; D006801:Humans; D007473:Ion Channels / Q000502:physiology; D008297:Male; D008854:Microscopy, Electron; D009419:Nerve Tissue Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D055495:Neurogenesis; D009474:Neurons / Q000166:cytology* / Q000502:physiology; D018408:Patch-Clamp Techniques; D018874:Spheroids, Cellular; D055815:Young Adult
chemical_list: D003470:Culture Media; D007473:Ion Channels; D009419:Nerve Tissue Proteins