Effect of ischemic culture conditions on the survival and differentiation of porcine dental pulp-derived cells

Although differentiated and undifferentiated cells can be exposed to ischemic conditions in cases of injury or inflammation, the effects of ischemia on cell survival and differentiation have not been well characterized. Here, we characterize the response of porcine dental pulp-derived cells (pDPCs) to culture conditions that approximate ischemia. Dental pulp is often exposed to ischemia due to narrow vascular openings in the tooth, which may affect the differentiation status of pDPCs. In this study, we investigated the influence of various ischemic conditions on differentiation-induced and non-induced pDPCs. To understand the character of cells used in this study, reported cell surface markers for dental pulp stem cells were investigated. pDPCs were CD90(low), CD105(+), and alpha-smooth muscle actin positive and showed osteogenic/chondrogenic differentiation potential. Anoxia was the most detrimental factor to cell viability, whereas hypoxia did not significantly affect survival. Glucose concentrations had a significant, mechanism-dependent effect on cell death. The presence of glucose correlated with caspase-dependent cell death, whereas the absence of glucose was linked to caspase-independent cell death. In contrast, differentiation status (i.e., induced versus non-induced pDPCs) did not affect the degree or mechanism of cell death. Finding depletion of specific markers by reverse transcription-polymerase chain reaction in both induced and non-induced cells suggests that the cells are de-differentiating under anoxia. Non-induced pDPCs were susceptible to anoxic induction of Oct-4, Sox-2, and hypoxia inducible factor-2 alpha, while these genes did not change in induced pDPCs. Re-differentiation analysis revealed that the surviving cells from non-induced pDPCs showed twofold higher alkaline phosphatase activity as compared with induced pDPCs, which suggest greater plasticity among the surviving fraction of non-induced pDPCs. These data showed that the ischemic conditions have similar detrimental influence on both undifferentiated and differentiated pDPCs, and affect differentiation status of pDPCs. Furthermore, ischemic conditions may influence the plasticity of undifferentiated pDPCs.