Abstract
Dental pulp stem cells (DPSCs) are a good source for tissue regeneration, however, the number of DPSCs in the pulp tissue is limited. Cell propagation is essential for tissue engineering using DPSCs and the cell culture conditions may affect the properties of DPSCs. The purpose of this study was to analyze the effect of cell culture condition, especially dense culture condition, on the property and differentiation pathway of DPSCs. We cultured DPSCs under sparse (sDPSCs; 5 × 103 cells/cm2) or dense (dDPSCs; 1 × 105 cells/cm2) conditions for 4 days and compared their properties. The populations of CD73+ and CD105+ cells were significantly decreased in dDPSCs. Both groups showed multi-differentiation potential, but mineralized nodule formation was enhanced in dDPSCs. The phosphorylation of focal adhesion kinase (FAK) and phosphoinositide 3-kinase (PI3K) proteins was promoted in dDPSCs, and alkaline phosphatase (ALP) mRNA expression in dDPSCs was abolished in the presence of pan-PI3K and FAK inhibitors. dDPSCs implanted into mouse bone cavities induced more mineralized tissue formation than sDPSCs and control. These findings indicate that dense culture conditions modified the properties of DPSCs and gave rise to osteogenic-lineage commitment via integrin signaling and suggest that dense culture conditions favor the propagation of DPSCs to be used for mineralized tissue regeneration.
Highlights
Mesenchymal stem cells (MSCs) derived from various mesenchymal tissues and organs are thought to be a good source for tissue engineering and regenerative medicine[1,2]
We here revealed that dental pulp stem cells (DPSCs) cultured under dense conditions exhibited reductions of CD73+ and CD105+ populations but maintained their multi-differentiation potential and acquired higher osteogenic differentiation potential in vitro and elevated bone-like tissue-forming ability in vivo
Our findings appear to support the notion that dense culture conditions promote the generation of cells committed to osteogenic differentiation from DPSCs
Summary
Mesenchymal stem cells (MSCs) derived from various mesenchymal tissues and organs are thought to be a good source for tissue engineering and regenerative medicine[1,2]. Dental pulp tissue contains dental pulp stem cells (DPSCs), which are undifferentiated neural crest-derived MSCs3. One of the advantages of DPSCs as a source for regenerative medicine is that the dental pulp tissue can be obtained from premolars planned to be extracted for orthodontic reasons or unfunctional/unnecessary wisdom teeth and supernumerary teeth, which are usually abrogated as waste[1]. Confluent culture conditions modify the properties of bone marrow stem cells (BMSCs), limiting their capacities to differentiate into www.nature.com/scientificreports/. We hypothesized that the density at which DPSCs are cultured influences their differentiation pathway, and evaluated the effects of sparse and dense cell culture conditions on their mesenchymal stem cell marker expression, proliferation, and capacity to differentiate into multiple lineages. We investigated the effects of cell culture conditions on their commitment to mineralized tissue-forming cells
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