The Role of C5a in Odontogenic Differentiation of Dental Pulp Stem Cells in Normoxia and Hypoxia

Regenerative endodontic procedures are treatment choices for young permanent immature teeth with chronic inflammation or necrosis of pulp tissue. Alterations in microenvironment will alter phenotypes of dental pulp stem cells (DPSCs) in pulp. The complement active fragment C5a is known to induce the recruitment of pulp progenitor cells to the injured site and we recently identified its critical roles in pulp fibroblast-mediated nerve regeneration under inflammatory injury. The role of C5a in the differentiation of DPSCs is unknown, especially under oxygen-deprived condition. The aim of this study is to determine the role of C5a in the odontogenic differentiation of DPSCs both in normoxia and hypoxia. Human DPSCs were isolated from healthy molars by the explant outgrown method. DPSCs were cultured in regular media and osteogenic media and treated with C5a antagonist-W54011 under normal and hypoxia conditions. Our PCR and immunohistochemistry data demonstrate that C5a plays a positive role in odontogenic differentiation of DPSCs. C5aR inhibition resulted in a significant decrease in odontogenic differentiation genes such as DMP1, ON, RUNX2, DSPP compared to the control. The hypoxic condition reversed this effect. Our results demonstrate that C5a constitutes a positive regulator of the odontogenic DPSC differentiation in normoxia. Under hypoxia, C5a exerts a negative function of DPSC differentiation. Taken together, we identified that C5a as a key in initial signal to control odontogenic DPSC differentiation providing the mechanism for potential therapeutic interventions of dentin repair and vital tooth preservation in evaluation of clinical outcome.