Induction of regulatory T cells and prolongation of survival of fully allogeneic cardiac grafts by administration of Tokishakuyaku-san in mice

Background: The Japanese herbal medicine Tokishakuyaku-san (TJ-23) has been used to treat neurodegenerative, immune, and respiratory tract diseases, as well as many gynecologic disorders, with few adverse effects. This study investigated the effect of TJ-23 on alloimmune responses in a murine model of cardiac allograft transplantation. Methods: CBA mice underwent transplantation of a C57BL/6 heart and received oral administration of 2 g/kg per day of TJ-23 or 1 of 16 other commonly used Japanese herbal medicines from the day of transplantation until 7 days afterward. An adoptive transfer study was conducted to determine whether regulatory cells were generated. Histologic and cell proliferation studies, cytokine measurements, and flow cytometry analyses were also performed. Results: Of the 17 herbal medicines studied, only TJ-23, given in a dose of 2 g/kg per day, induced significantly prolonged allograft survival (median survival time [MST], >100 days). TJ-23 also suppressed proliferation of splenocytes and production of interleukin-2, interleukin-6, and interferon-γ. Adoptive transfer of either whole splenocytes or CD4 ⁺ or CD4 ⁺ CD25 ⁺ cells from TJ-23-treated allograft recipients resulted in indefinite survival of allografts in naive secondary recipients (MST >100 days). Flow cytometry studies showed that the CD4 ⁺ CD25 ⁺ forkhead/winged-helix (FOXP3) ⁺ regulatory cell population was increased in transplant recipients given TJ-23. Conclusion: TJ-23 induced hyporesponsiveness to fully allogeneic cardiac allografts and generated CD4 ⁺ CD25 ⁺ regulatory cells in our model.