Rapid differentiation of human dental pulp stem cells to neuron-like cells by high K⁺ stimulation

Abstract

As human-origin cells, human dental pulp stem cells (hDPSCs) are thought to be potentially useful for biological and medical experiments. They are easily obtained from lost primary teeth or extracted wisdom teeth, and they are mesenchymal stem cells that are known to differentiate into osteoblasts, chondrocytes, and adipocytes. Although hDPSCs originate from neural crest cells, it is difficult to induce hDPSCs to differentiate into neuron-like cells. To facilitate their differentiation into neuron-like cells, we evaluated various differentiation conditions. Activation of K + channels is thought to regulate the intracellular Ca 2+ concentration, allowing for manipulation of the cell cycle to induce the differentiation of hDPSCs. Therefore, in addition to a conventional neural cell differentiation protocol, we activated K + channels in hDPSCs. Immunocyto­chemistry and real-time PCR revealed that applying a combination of 3 stimuli (high K + solution, epigenetic reprogramming solution, and neural differentiation solution) to hDPSCs increased their expression of neuronal markers, such as β3-tubulin, postsynaptic density protein 95, and nestin within 5 days, which led to their rapid differentiation into neuron-like cells. Our findings indicate that epigenetic reprogramming along with cell cycle regulation by stimulation with high K + accelerated the differentiation of hDPSCs into neuron-like cells. Therefore, hDPSCs can be used in various ways as neuron-like cells by manipulating their cell cycle.

Peripheral-neuron-like properties of differentiated human dental pulp stem cells (hDPSCs)

“…In our experiments, βIII-tubulin-positive processes were observed in hDPSCs after seven days in differentiation media, and bipolar type neuron-like cells with elongated neurite-like processes were observed after fourteen days ( Fig 1 ). Whereas differentiated and undifferentiated hDPSCs also expresses a neuronal stem cell marker, nestin, and an astrocyte marker, glial fibrillary acidic protein (GFAP) ( S1 Fig and also reported from other laboratories [ 37 , 50 ]), we have demonstrated that the mRNA expression level of GFAP is much lower than that of βIII-tubulin in undifferentiated and differentiated hDPSC [ 51 ]. Therefore, βIII-tubulin predominates over GFAP in hDPSCs.…”

Influence of gap junctions upon Ca²⁺ propagation from stimulated keratinocytes to DRG neurons

“…The present question is how we can obtain human free nerve endings. One possible answer is the use of human dental pulp stem cells [ 37 , 38 ]. If an in vitro model for a human free nerve ending model in the skin is established, the development of analgesics will advance considerably.…”

Dorsolateral prefrontal cortex sensing analgesia

“…Human epidermal keratinocytes and dermal fibroblasts can be easily obtained as commercially available cells, whereas human neurons are difficult to collect. Oka and Ito have attempted to prepare neurons by differentiating human dental pulp stem cells (hDPSC) [ 66 , 67 ] for this purpose. In particular, Arimura and colleagues compared neurons originating from hDPSCs and rat neurons isolated from the dorsal root ganglion (DRG) [ 67 ].…”