Hypoxia upregulates the expression of the pluripotency markers in the stem cells from human deciduous teeth

Stefanie Bressan Werle, Pedro Chagastelles, Patricia Pranke, Luciano Casagrande

Cultivation under hypoxia promotes different responses in the mesenchymal stem cells and it has been producing promising results for clinical applications. Pulp tissue from deciduous teeth is a source of stem cells which has a high proliferative potential but this is usually discarded. This study has evaluated the effects of hypoxia on proliferation, apoptosis, and the expression of the pluripotency-related genes of the stem cells from human exfoliated deciduous teeth (SHED). The cells were isolated from dental pulp (n = 5) and characterized as mesenchymal stem cells, in accordance with the International Society for Cell Therapy. The cells were cultivated under hypoxia (3% oxygen) and compared to the normoxia cells (21% oxygen). The proliferation rate was evaluated by the Ki67 antibody for up to 7 days, while the metabolic activity was measured by the wst-8 assay for up to 14 days. The apoptotic cells were analyzed by Annexin V and propidium iodide staining at 24 h and 4 and 7 days. The expression of the pluripotent genes (OCT4, SOX2, and NANOG) was quantified by qPCR after 24 h, or 7 days, when cultivated under hypoxia or normoxia. No differences in the metabolic activity, the proliferation rate, and the apoptosis of SHED when cultivated under hypoxia or normoxia (p > 0.05) were observed. The expression of the pluripotent genes was significantly higher after 24 h and 7 days of the cells that were exposed to hypoxia (p < 0.01). These findings have indicated an increase of the pluripotency-related genes within 7 days as being the main advantage of SHED culture under hypoxia. Hypoxia culture may help maintain the quiescent state of the SHED, which could be advantageous for their future clinical applications.