Calcium hydroxide has been extensively and steadily used for direct pulp capping in modern clinical dentistry. As it was known to have potential to induce hard tissue repair, this chemical has been applied to the exposed dental pulp and the hard tissue is expected to be regenerated above the pulp. During the reparative process of exposed pulp, primary odontoblasts that were lost as a result of extensive damage are replaced with newly differentiated odontoblast-like cells. This process is known to follow the sequential steps of proliferation, migration, and differentiation of progenitor cells. This research will examine the relationship between calcium hydroxide and the recruitment, proliferation, and mineralization of postnatal dental stem cells, obtained from an immature dental tissue of beagle dogs. Immunocytochemical staining and reverse transcriptase-polymerase chain reaction were used to identify the putative stem cell markers. Immunoblot analysis, wound healing assay, cell migration assay, and alizarin red staining were used to evaluate proliferation, migration, and mineralization capacity of the calcium hydroxide-treated stem cells. As an in vivo study, a combination of calcium hydroxide and autologous dental pulp stem cells (DPSCs) was applied for the treatment of intentionally created tooth defects on the premolars and the molars in beagle dogs to observe dentin regeneration. Ex vivo expanded DPSCs and periodontal ligament stem cells expressed STRO-1 and CD146, the mesenchymal stem cell markers. It was evident that calcium hydroxide increased recruitment, migration, proliferation, and mineralization of the DPSCs and periodontal ligament stem cells. Such results are valuable for future availability of DPSCs, which are recently focused as the stem cell reservoir for regeneration of dentin upon tooth injury, as well as for elucidation of the role of calcium hydroxide in pulp capping therapy.