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

TLDR

The study aimed to demonstrate that hDPSCs from third molar pulp retain multilineage differentiation after cryopreservation. hDPSCs were isolated, cryopreserved, thawed, and then cultured in neurogenic, osteogenic/odontogenic, adipogenic, myogenic, and chondrogenic media, with proliferation, STRO‑1 expression, and lineage marker analysis performed, and results compared to an NIH‑derived strain. The cryopreserved hDPSCs proliferated for at least 25 passages, expressed STRO‑1, and differentiated into all five lineages, with the primary culture favoring myogenesis and the NIH strain favoring odontogenic/osteogenic and chondrogenic differentiation, supporting third molar pulp as a viable multipotent stem cell source.

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.