Introduction: The purpose of this study was to assess the significant proliferation of dental pulp-derived stem cells in vitro from rats with the systemic administration of immunosuppressant in subcutis. There must be a sufficient number of stem cells for tooth regeneration. However, number of mesenchymal stem cells in the dental pulp tissue is a small. Then, the proliferation of stem cells must be accelerated for hard tissue formation. The subcutaneous injection of the immunosuppressant would enhance the hard tissue forming ability of dental pulp cells of rat. It was hypothesized in this study that differentiation of stem cells into blasts would be effectively promoted by suppression of the systemic immune response. Materials and methods: The dental pulp cells of rats with immunosuppressant injection subcutaneously were cultured with or without addition of the immunosuppressant in the medium containing dexamethasone for calcified nodule formation. Ca2+ by decalcification of calcified nodules were quantitatively analysed. Statistical comparisons between the quantities of Ca2+ were performed using two-way unrepeated ANOVA followed by post hoc analysis with Tukey-Kramer’s test. Differences of p < 0.01 were considered significant. Results: The proliferation and differentiation of stem cells among dental pulp cells was inhibited by the presence of immunosuppressive agents in the culture medium. However, stem cells obtained from rats after systemic administration of an immunosuppressive agent exhibited a high ability to form calcified nodules. Conclusions: To promote proliferation and differentiation of stem cells, systemic administration of an immunosuppressant to individuals prior to harvesting stem cells would be recommended.