Multilineage Differentiation Potential of Stem Cells Derived from Human Dental Pulp after Cryopreservation

Abstract

The current study aimed to prove that human dental pulp stem cells (hDPSCs) isolated from the pulp of third molars can show multilineage differentiation after cryopreservation. First, hDPSC were isolated via enzymatic procedures, and frozen in liquid nitrogen until use. After defrosting, cells were analyzed for proliferative potential and the expression of the stem cell marker STRO-1. Subsequently, cells were cultured in neurogenic, osteogenic/odontogenic, adipogenic, myogenic, and chondrogenic inductive media, and analyzed on basis of morphology, immunohistochemistry, and reverse transcriptase-polymerase chain reaction (RT-PCR) for specific marker genes. All data were replicated, and the results of the primary cells were compared to similar tests with an additional primary dental pulp stem cell strain, obtained from the National Institutes of Health (NIH). Results showed that our cell population could be maintained for at least 25 passages. The existence of stem/ progenitor cells in both cell strains was proven by the STRO-1 staining. Under the influence of the 5 different media, both cell strains were capable to advance into all 5 differentiation pathways. Still differences between both strains were found. In general, our primary culture performed better in myogenic differentiation, while the externally obtained cells were superior in the odontogenic/osteogenic and chondrogenic differentiation pathways. In conclusion, the pulp tissue of the third molar may serve as a suitable source of multipotent stem cells for future tissue engineering strategies and cell-based therapies, even after cryopreservation.

Osteoblastic/Cementoblastic and Neural Differentiation of Dental Stem Cells and Their Applications to Tissue Engineering and Regenerative Medicine

“…24,25 To date, in addition to osteogenic differentiation, several differentiation studies of dental stem cells have reported on their capacity to differentiate into chondrogenic, adipogenic, neural, and endothelial cell lineages. 23,26,32,33 For instance, adipogenic differentiation was induced by the addition of 0.5 mM isobutyl-methylxanthine, 1 mM dexamethasone, 10 mM insulin, 200 mM indomethacin, and 50 mg/mL of gentamicin into alpha minimum essential medium (a-MEM) containing 10% fetal calf serum (FCS). 32 Such adipogenic differentiation capacity was also reported in PDLSC differentiation studies.…”

“…23,26,32,33 For instance, adipogenic differentiation was induced by the addition of 0.5 mM isobutyl-methylxanthine, 1 mM dexamethasone, 10 mM insulin, 200 mM indomethacin, and 50 mg/mL of gentamicin into alpha minimum essential medium (a-MEM) containing 10% fetal calf serum (FCS). 32 Such adipogenic differentiation capacity was also reported in PDLSC differentiation studies. 23,26 In addition, despite that chondrogenic differentiation activity was relatively lower in comparison with BMSC, chondrogenic differentiation of dental stem cells could be achieved by the micromass culture of DPSC in the presence of 10% FCS, 200 mg/mL ascorbic acid, and 10 ng/ mL of TGF-b3 after 6 weeks.…”

“…34 In addition to skeletal lineage differentiation of dental stem cells, different lineage differentiations such as neural and endothelial differentiation could be induced under certain culture conditions. 32,35 Neural differentiation was induced by the culture in the presence of 10 mM b-mercaptoethanol, 2% dimethyl sulfoxide, and 200 mM butylated hydroxyanisole without serum. 32 Nakasima et al reported in vitro and in vivo endothelial (endothelial cell lineage) differentiation capacity of DPSC.…”

“…32,35 Neural differentiation was induced by the culture in the presence of 10 mM b-mercaptoethanol, 2% dimethyl sulfoxide, and 200 mM butylated hydroxyanisole without serum. 32 Nakasima et al reported in vitro and in vivo endothelial (endothelial cell lineage) differentiation capacity of DPSC.…”

Dental pulp can be a good candidate for nerve grafting in a xeno-graft model

“…This anatomical structure may 6 be applicable for nerve conduit. Besides, dental pulp contains affluent growth factors as well as mesenchymal progenitor cells (Nosrat et al, 2001), and dental pulp stem cell is one of the topics in the regenerative medicine (Zhang et al, 2006). However, there is no trial thus far to use the dental pulp as a nerve scaffold.…”