Abstract
BackgroundThe concept of establishing a dental stem cell (DSC) bank for oral and maxillofacial regeneration has become of great interest but it remains at a primitive stage. The routine application of serum-containing conditions for human DSC (hDSC) culture is in great controversy considering that the animal-originated serum can cause serious ethical concerns and lead to increasingly irrelevant variables, errors, and poor repeatability of experiment results. Thus, this study aimed to establish a safe, stable and efficient hDSC serum-free culturing system for future DSC bank usage.MethodsDental pulp stem cells (DPSCs) from human permanent tooth pulp were isolated, expanded, passaged, and divided into two groups according to their culture conditions: group 1 was the serum-containing medium (SCM) group; and group 2 was the serum-free Essential 8 medium (E8) group. DPSCs were characterized first, followed by cell proliferation, pluripotency, and migration study in SCM and E8 medium.ResultsHuman DPSCs (hDPSCs) in E8 medium demonstrated greater proliferation, pluripotency, migration ability and less apoptosis. hDPSCs could be successfully induced to the adipogenic, osteogenic, neurogenic, and chondrogenic lineages in E8 group. Real-time polymerase chain reaction indicated that the expression of PPAR-γ, RUNX2, OCN and MAP-2 was higher in E8 group. ConclusionsCompared with serum-containing medium, E8 medium exhitibed higher ability in maintaining the cell proliferation, pluripotency, migration, and stability. This new serum-free culture environment might be applicable for hDSC culture in the future.
Highlights
The concept of establishing a dental stem cell (DSC) bank for oral and maxillofacial regeneration has become of great interest but it remains at a primitive stage
After cells reached a concentration of 80%, they were cultured under separate conditions: in the Essential 8 medium (E8) group, the cell suspensions were incubated at a cell density of 2 × 105 cells per culture plate in albumin-free E8 culture medium (Stemcell, USA) containing chemically defined and concentration-determined multiple factors; in the serum-containing medium (SCM) group, the cell suspensions were plated at the same cell density in normal medium containing Dulbecco’s modified Eagle’s medium (DMEM) and 5% fetal bovine serum (FBS) as a control
Changes in cell morphology Cells cultured in SCM proliferated sparsely in a single layer and demonstrated typical spindle and polygonal
Summary
The concept of establishing a dental stem cell (DSC) bank for oral and maxillofacial regeneration has become of great interest but it remains at a primitive stage. Increasing evidence has indicated that FBS is a controversial ingredient owing to its risk of transmitting prion, zoonotic, or viral infections, and the xenogeneic compounds can trigger host immune responses, which are a potential and significant hazard. Another important concern is the batch-to-batch variety of the quality and protein concentration [10,11,12]. Bonnamain et al [16] reported successful expansion of human stem cells in a chemically defined serum-free culture medium with added growth factors. Discoveries of signal pathways and multiple factor mechanisms in serum-free medium are showing steady progress, and a number of genes connected with the response have been unraveled, contributing to the use of hPSCs as engineering tools for therapeutic purposes [17,18,19,20,21]
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