Use of a 3D Floating Sphere Culture System to Maintain the Neural Crest-Related Properties of Human Dental Pulp Stem Cells

Abstract

Human dental pulp is considered an interesting source of adult stem cells, due to the low-invasive isolation procedures, high content of stem cells and its peculiar embryological origin from neural crest. Based on our previous findings, a dental pulp stem cells sub-population, enriched for the expression of STRO-1, c-Kit, and CD34, showed a higher neural commitment. However, their biological properties were compromised when cells were cultured in adherent standard conditions. The aim of this study was to evaluate the ability of three dimensional floating spheres to preserve embryological and biological properties of this sub-population. In addition, the expression of the inwardly rectifying potassium channel Kir4.1, Fas and FasL was investigated in 3D-sphere derived hDPSCs. Our data showed that 3D sphere-derived hDPSCs maintained their fibroblast-like morphology, preserved stemness markers expression and proliferative capability. The expression of neural crest markers and Kir4.1 was observed in undifferentiated hDPSCs, furthermore this culture system also preserved hDPSCs differentiation potential. The expression of Fas and FasL was observed in undifferentiated hDPSCs derived from sphere culture and, noteworthy, FasL was maintained even after the neurogenic commitment was reached, with a significantly higher expression compared to osteogenic and myogenic commitments. These data demonstrate that 3D sphere culture provides a favorable micro-environment for neural crest-derived hDPSCs to preserve their biological properties.

Regenerative potential of human dental pulp stem cells in the treatment of stress urinary incontinence: In vitro and in vivo study

“…An equal area was selected inside each band, and the mean of grey levels (in a 0‐256 scale) was calculated. Data were then normalized to values of background and of control actin band …”

Advances of tooth‐derived stem cells in neural diseases treatments and nerve tissue regeneration

“…Conditioned media from dental stem cells can enhance the growth rate of Schwann cells and induce the neurite outgrowth in vitro. 44,45 Additionally, inner neurotrophic factors such as BDNF, NT-3 and GDNF can promote the neural differentiation of DPSCs and SHEDs. However, among all the dental stem cells, SCAPs have shown the greatest potential for neurotrophic effects, indicating that SCAPs could be an optimal cell source for peripheral nerve repair.…”

“…Functional voltage-gated Na + channels are present in DPSCs when exposed to neuronal inducing conditions. 44,45 Additionally, inner neurotrophic factors such as BDNF, NT-3 and GDNF can promote the neural differentiation of DPSCs and SHEDs. The DPSCs and SHEDs differentiated cells showed comparable functional neural activities, indicating that both adult and adolescent teeth could be considered as a cell-based therapy source for neural diseases treatments.…”

Human Dental Pulp Stem Cells: recent findings and current research

“…Bearing in mind that cell arrangement, growth factors and biological scaffolds in total promote induced localized proliferation, researchers currently employ the cell bead and cell sheet method in which layer by layer, compact and spongy bone tissue constructs containing vascular networks are printed [9]. Future application of STRO-1+cK-it+CD34+ DPSCs is based on their ability to form 3D sphere system, capacity to preserve immunological expression of STRO-1+cKit+CD34+ and expression of late neuronal differentiation markers, but also on maintaining capacity for osteogenic differentiation [47]. Specifically, STRO-1+cKit+FLK-1+CD34+ cells were reported to have the highest growth potential and neurogenic commitment, while STRO-1+cK-it+CD34+ alone express significantly more low affin- [11] general: STRO-1, CD29, CD44, CD73, CD90, CD105, CD146, CD166, CD271 hematopoietic: CD34 and CD117 stemness: OCT-3/4 and NANOG [25] CD13, CD29, CD105, CD146, CD166 [1] CD90, CD73, CD34 [49] STRO-1, c-Kit, CD29, CD34, CD44, CD73, CD105, CD146 [47] STRO-1, c-Kit, CD34 [15] CD90, CD73, CD34 ity nerve growth factor receptor (LNGFR) [47,48].…”

“…Future application of STRO-1+cK-it+CD34+ DPSCs is based on their ability to form 3D sphere system, capacity to preserve immunological expression of STRO-1+cKit+CD34+ and expression of late neuronal differentiation markers, but also on maintaining capacity for osteogenic differentiation [47]. Specifically, STRO-1+cKit+FLK-1+CD34+ cells were reported to have the highest growth potential and neurogenic commitment, while STRO-1+cK-it+CD34+ alone express significantly more low affin- [11] general: STRO-1, CD29, CD44, CD73, CD90, CD105, CD146, CD166, CD271 hematopoietic: CD34 and CD117 stemness: OCT-3/4 and NANOG [25] CD13, CD29, CD105, CD146, CD166 [1] CD90, CD73, CD34 [49] STRO-1, c-Kit, CD29, CD34, CD44, CD73, CD105, CD146 [47] STRO-1, c-Kit, CD34 [15] CD90, CD73, CD34 ity nerve growth factor receptor (LNGFR) [47,48]. Therefore, mainly STRO-1+ DPSCs show generally firmer tendency for neurogenic commitment rather than other pulp cells expressing common mesenchymal markers [49,50].…”