Exosome-based Cell Homing and Lineage-Specific Differentiation Strategies for Dental Pulp Regeneration

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[PROJECT SUMMARY/ABSTRACT] Current endodontic therapy for dental caries, which are one of the most prevalent infectious diseases in the world, is a procedure for replacing the vital pulp with synthetic pulp-capping materials. Pulpless teeth can lose their functions to sense environmental changes and maintain dentin regeneration, and the synthetic materials have several disadvantages such as bacterial leakage into the dental pulp, poor cohesive strength, discoloration of tooth, and long setting time. As an alternative, vital pulp therapy (VPT), which is defined as a restorative dental treatment that aims to preserve and maintain pulp tissue, is beneficial for young patients who have high healing capacity for pulp regeneration. Potential for successful VPT and pulp regeneration is increasing due to the knowledge of mesenchymal stem cells (MSCs) that can differentiate into specialized cells. However, the transplantation of MSCs incurs high costs and risks associated with the ex vivo cell expansion. Consequently, a cell homing strategy which recruits endogenous dental pulp stem cells (DPSCs) is the effective approach in endodontics. Recently, exosomes have attracted attention due to their great potential to promote intercellular communication leading to enhanced cell recruitment, differentiation to specific cell lineage, and tissue regeneration. In particular, conditioned medium or exosomes cultured under lineage-specific differentiation have a great potential for angiogenesis and odontogenesis for pulp regeneration. The long-term goal is to develop a pulp capping material system for vital pulp therapy of human dental pulp. The overall objectives for this application are (1) to elucidate the therapeutic potential of characterized exosomes as a chemoattractant to stimulate DPSC migration and pulp-like differentiation and (2) to determine their in vivo effect of pulp regeneration in a rabbit partial pulpotomy model. Our central hypothesis is that exosomes will stimulate dental pulp regeneration by promoting DPSC chemotaxis and lineage-specific differentiation. Exosomes will be isolated from rabbit DPSCs cultured under growth or lineage-specific differentiation conditions (odontogenesis or angiogenesis) and will be encapsulated in injectable hydrogel (F-127/hyaluronic acid) which has temperature-sensitive gelation behavior at body temperature. Specific aims in this proposed study are (1) characterize DPSC-Exos and determine the effect of DPSC-Exos on in vitro cell homing and lineage-specific differentiation and (2) evaluate the in vivo effects of DPSC-Exos on pulp regeneration in a rabbit partial pulpotomy. At the completion of the proposed R03 project, our expected outcomes are to define the therapeutic potential of DPSC-Exos for dental pulp regeneration and to identify miRNAs that may regulate cell homing and pulp tissue formation. These results will have a very important positive impact by providing preliminary data for our future R01 grant application in which we plan to synthesize miRNAs loaded in engineered exosomes, thereby replacing DPSC culture as a potential means of exosome production.