Effects And Mechanisms Of Calcineurin Inhibitor On Development Of Murine Th17 Cells

Effects and Mechanisms of Calcineurin Inhibitor on Development of Murine Thl7 cellsObjectiveTransplantation is the optimal treatment for many end-stage diseases in solid organs Despite progress in immuno suppression, allograft rejection is still a major challenge tc long-term survival of recipients and grafts. The immuno logic role of T helper type 17 cells a recently described CD4+ T-helper cell subset, has been implicated in a range o: autoimmune diseases and inflammatory responses previously believed to be mediated b) Thl. Studies have suggested the presence of both Thl and Thl7 cells during allograf rejection: along with Thl cells, Thl7 cells may also mediate allograft rejection, and neithe: subset is sufficient for the occurrence of rejection. When Thl cells are inhibited, Thl7 cells may have a more important role in allograft rejection. To prevent allograft rejection, it h necessary to understand the factors that affect development of T-cell subsets. Calcineurir inhibitor is a T-cell targetted immunosuppression drug that is widely used to preven allograft rejection, it exerts immunosuppression effects by inhibition of T-cell activation, ir particular, Thl cell. However, its pharmacologic effects on Thl7 cells have not been fiill> elucidated. To evaluate whether calcineurin inhibitor inhibits the activities of Thl7 cells cells were differentiated under Thl7 polarized conditions in the presence of increasing amounts of the calcineurin inhibitor such as tacrolimus and cyclosporine A. To establish heterotopic cardiac transplantation model in mice to provide an effective means fo] studying the effect and mechanism of calcineurin inhibitor on Thl7 cells. Besides, tc investigate the role of Thl 7 cells and their cytokines in cardiac allogrft rejection in mice. MethodsMurine naive CD4+CD25~ T cells were stimulated with plate-bound anti-CD3 and anti-CD28; then were induced by the cytokines such as TGFpl and IL-6, etc. Calcineurin inhibitor such as tacrolimus and cyclosporine A was added to the cells at the desired time point and desired concentrations. The subsets of CD4+Thl7 cells were examined by flow cytometry. The expression of IL-17 mRNA was assayed by RT-PCR and Western Blot. The heterotopic cardiac transplantation models were divided into 4 groups: isograft group, acute rejection group, CsA group and FK506 group. Mean survival time was measured. The mouse hearts in each group were harvested on the day 1, 2, 4, 6, and 8 post transplantation. The mRNA expression of IL-17 and IFNy was detected by RT-PCR.ResultsCalcineurin inhibitor inhibits the differentiation and proliferation of Thl7 cells in a dose-dependent manner. The differences among the groups are considered significant(P<0.05). In the acute rejection group, RT-PCR revealed that IL-17 mRNA was expressed earlier than IFNy, its expression was detected on the second day after transplantation, the expression quantity reached the peak on the postoperative day 4, the gradually reduced, and on the day 6 and 8, no expression was detectable.ConclusionThl7 cells may play an important role in the development of allograft rejection, and IL-17 could serve as a predictive parameter for allograft rejection in the future. The present study investigated the effects of calcineurin inhibitor on Thl7 cells. Inhibition o: differentiation and proliferation of Thl7 cells was dose-dependent. Experssion of IL-1* mRNA was also inhibited. A new mechanism is necessary to explain this phenomenon Gomez-Rodriguez, et al. Observed that expression of the ill7a gene in Thl7 cells is particularly sensitive to the strength ofT-cell receptor signaling, requiring full activation o: Ca2+ mediated pathways in addition to cytokine signals requried for induction and activation of retinoic acid-related orphan receptor-yt and signal transducer and activator o: transcription-3. Expression ol IL-17A was rescued by pharmacologically induced Ca2" influx or activated NFATc; decreased T-cell receptor stimulation or calcineurin inhibitior preferentially reduced IL-17A expression. Expression of IL-17Ais specifically coupled tc T-cell receptor signaling via calcineurin-inhibited regulation of NFATc. In summary calcineurin inhibitor inhibits calcineurin in Thl7 cells, blocks dephosphorylation anc translocation of NFATc, and inhibits transcription of the ill7a gene, reducing expressior IL-17A and alleviating the process of inflammation, which in turn contributes tc suppression of allograft rejection. Fingding of the present study shed new light on the potential immuno suppres sive mechanisms of calcineurin inhibitor and presents anc experimental foundation and theoretical basis for clinical immunosupprission protocols.