The Authors' Reply.

The Authors’ Reply Byung Ha Chung, and Chul Woo Yang Th17 is the most recently discovered CD4 + T cell subset, and accumulating evidence show that Th17 cells are involved in driving immune processes previously thought to be exclusively Th1mediated in various immune diseases, including allograft rejection in kidney transplantation (KT). However, tacrolimus (Tac), the main immunosuppressant in solid organ transplantation, showed limited suppressive impacts on Th17-related immune responses in vitro and in vivo. In this regard, we were interested in the strategy to effectively regulate Th17 cells in KT recipients under Tac-based immune suppression. At first, we published a study which showed that the mTOR inhibitor, rapamycin, had better suppressive effects on Th17 cells than Tac. However, it is well known that, although the in vitro results of the mTOR inhibitor are promising, its immune suppressive effects in terms of prevention of acute rejection in clinical practice is inferior to that of Tac. Therefore, we switched our strategy to compensate the effects of Tac rather than conversion of Tac to another drug. Therefore, in the next study, we focused on the modulating effects of vitamin D on the mTOR pathway. Indeed, the active form of vitamin D can suppress mTOR directly or through PI3k/akt, S6k and STAT3 molecule, which result in suppression of the Th17 pathway. However, as indicated in the recent article by Aly et al, the dose or concentration of 25(OH)D3 or 1,25(OH)D3 that show significant immune regulatory effects of vitamin D is an important issue. However, in our study, the specific conditions that were applied should be considered; both in vitro and in vivo conditions included not only vitamin D but also Tac. Although the effects of Tac on Th17 is limited, combined use of vitamin D and Tac can show a synergistic impact on the suppression of the mTOR pathway and Th17 pathway as shown in our study, as well as in another study on autoimmune disease. In our additional in vitro experiments, Tac + mycophenolate mofetil (MMF) did not show significant suppressive effects on the appearance of IL-17–positive T cells. However, the addition of 1,25(OH)2D3 to the Tac + MMF condition caused a significant reduction in the percentage of these cells. IL-17 or IL-22 levels in the culture supernatant also showed significant decrease after the addition of 1,25(OH)2D3. Therefore, it is possible that even the dose of vitamin D, which may be considered suboptimal for immune modulation when used alone, may effectively regulate Th17 in clinical situations under Tac-based immunosuppression. In addition, clinical correlation was shown with between low concentrations of 25(OH)D3 and immunologic events in our previous study and in another study by Kwon et al, in KT recipients under maintenance immunosuppression. Therefore, we concluded that vitamin D by itself may not have significant immunosuppressive actions; however, when it is added on to another stronger immune suppressant such as Tac, it can show some significant effects, which is the main theme of our study. Considering this point, vitamin D is an immune modulator rather than an ineffective immune suppressor as indicated by Aly et al. Nevertheless, further studies including the measurement of both serum 25(OH)D3 and 1,25(OH)D3 levels or examination of vitamin D receptor expression on T cells before and after calcitriol therapy may be necessary. Also, the above-mentioned data cannot give us more useful information on the complex correlations of Tac, vitamin D, and Th17. Unfortunately, we could not secure enough serum and PBMC in our study; hence we could not examine those parameters. In addition, a more delicate in vitro model may be necessary to prove the synergistic effect of Tac and vitamin D on Th17 cells. Lastly, to clarify all the above issues, a randomized controlled trial is required.