Dental Pulp of the Third Molar: A New Source of Pluripotent-like Stem Cells

Abstract

SummaryDental pulp is particularly interesting in regenerative medicine because of the accessibility and differentiation potential of the tissue. Dental pulp has an early developmental origin with multi-lineage differentiation potential as a result of its development during childhood and adolescence. However, no study has previously identified the presence of stem cell populations with embryonic-like phenotypes in human dental pulp from the third molar. In the present work, we describe a new population of dental pulp pluripotent-like stem cells (DPPSCs) that were isolated by culture in medium containing LIF, EGF and PDGF. , and they show genetic stability in vitro based on genomic analysis with a newly described CGH technique. Interestingly, DPPSCs were able to form both embryoid-body-like structures (EBs) in vitro and teratoma-like structures that contained tissues derived from all three embryonic germ layers when injected in nude mice. We examined the capacity of DPPSCs to differentiate in vitro into tissues that have similar characteristics to mesoderm, endoderm and ectoderm layers in both 2D and 3D cultures. We performed a comparative RT-PCR analysis of GATA4, GATA6, MIXL1, NANOG, OCT3/4, SOX1 and SOX2 to determine the degree of similarity between DPPSCs, EBs and human induced pluripotent stem cells (hIPSCs). Our analysis revealed that DPPSCs, hIPSC and EBs have the same gene expression profile. Because DPPSCs can be derived from healthy human molars from patients of different sexes and ages, they represent an easily accessible source of stem cells, which opens a range of new possibilities for regenerative medicine.

Notch/Wnt cross-signalling regulates stemness of dental pulp stem cells through expression of neural crest and core pluripotency factors

“…These cells are easily www.ecmjournal.org accessible in both young and aged patients, possess a significant capacity for in vitro expansion and have non-tumourigenic phenotypes. Furthermore, evidence shows that DPSCs have a greater multilineage differentiation potential compared to other tissue-specific stem cells (Atari et al, 2012;Kerkis et al, 2006;Rosa et al, 2016). It is illustrative of the growing acknowledgement of the stemness potential of DPSCs that some authors have referred to some subpopulations of these cells as dental pulp pluripotent stem cells or DPPSCs (Atari et al, 2012).…”

“…This very important finding proved that DPSC exhaustion by continuous exposure to FBS could be at least partially prevented by transiently boosting the activity of Notch/Wnt pathways, by means of small molecules and/or recombinant proteins, which would enhance stemness traits of DPSC cultures just 48 h before use for differentiation or cell transplant. It would also be very interesting to verify whether such preconditioning would also be effective for other non-mesenchymal cell lineages of interest, such as Schwann cells (Martens et al, 2014), neuron-like cells (Gervois et al, 2015) and hepatocyte-like cells (Atari et al, 2012). Finally, the adoption of this preconditioning strategy could also be applicable to differentiation protocols that do not rely on FBS (Eubanks et al, 2014;Xiao and Tsutsui, 2013), allowing for a faster translation to clinical therapy.…”

“…Other stem cell types with similar properties may be obtained from gingival and periodontal tissues (Abe et al, 2012;Huang et al, 2009;Ibarretxe et al, 2012;Lima et al, 2017;Liu et al, 2015;Petrovic and Stefanovic, 2009) and even from periodontal inflammatory lesions, such as human periapical cysts (Marrelli et al, 2013;Marrelli et al, 2015;. Similarly to NC-derived cells, dental stem cells are characterised by the expression of neural crest factors, such as Snail/Snai1, Slug/Snai2, Twist1, Hnk1, Pax3, Neurogenin2 and Sox10 (Kiraly et al, 2009;Schiraldi et al, 2012), and core factors including Oct4a, Sox2, Klf4, Lin28, stage specific embryonic antigen 3 antibody (Ssea1) and Nanog (Atari et al, 2012;Ferro et al, 2012;Janebodin et al, 2011;Kerkis et al, 2006;Rosa et al, 2016). Core factors are fundamental in maintaining stem cell pluripotency (Chambers and Tomlinson, 2009;Takahashi et al, 2007;Yu et al, 2007), thus suggesting that dental stem cells may present some superior features with www.ecmjournal.org V Uribe-Etxebarria et al…”

“…Notch/Wnt crosstalk regulates DPSC stemness respect to other multipotent stem cell populations of the adult human body (Atari et al, 2011;Atari et al, 2012;Rosa et al, 2016). This could be very relevant for cell therapy because stem cells from dental tissues are known to be easily accessible for extraction and well-tolerated upon grafting, due to their immunesuppressive properties (Pierdomenico et al, 2005).…”

“…Both pathways also play an important role in the emergence of the NC (Hari et al, 2012;Leung et al, 2016;Rogers et al, 2012;Stuhlmiller and Garcia-Castro, 2012). Dental stem cells present higher levels of core factors and Wnt/Notch activity compared to other mesenchymal stem cells in the adult body (Atari et al, 2012;Huang et al, 2009;Janebodin et al, 2011;Vasanthan et al, 2015). However, the role of these pathways in the maintenance of stemness and self-renewal in DPSCs is still unclear.…”