Background: FoxP3+ follicular regulatory T cells (Tfr) have been identified as the cell population controlling T follicular helper (Tfh) cells and B cells that are involved in effector immune responses against transplanted tissue. 
Materials: To understand the biology of Tfr cells in kidney transplant patients receiving tacrolimus based immunosuppression, we measured circulating (c)Tfh and cTfr cells in peripheral blood at 5-7 years after transplantation by flow cytometry. Of this cohort 26% (58/227) had been treated for rejection of which 16% was diagnosed as T cell mediated rejection (TCMR), 7% as chronic (c)ABMR and 3% as mixed rejection. Median time after anti-rejection therapy was 4.9 years (range 0.4–7 years). Age and gender matched heathy individuals served as controls.
Results: While the absolute numbers of cTfh cells were comparable between kidney recipients and healthy controls, the numbers for cTfr cells were 46% lower in immunosuppressed recipients (median 8.8×10⁶/mL vs. 16.2×10⁶/mL, p<0.001). More importantly, the ratio of Tfr/Tfh was decreased, indicating a disruption of the balance between cTfh and cTfr cells. This shifted balance was observed for both non-rejectors and rejectors. Next, the impact of a previous anti-rejection therapy history on cTfr cells. After pulse steroid anti-rejection therapy 29% (median) fewer cTfr cells were measured, after IvIG a 40% reduction was found and five years after alemtuzumab therapy hardly any cTfr cells were present (p<0.01 for all group vs non-rejectors). No association with tacrolimus trough levels was found. The Tfr/Tfh ratio correlated with kidney function (eGFR: rs=0.15, p=0.03).
Conclusions: Our work shows that anti-rejection therapy significantly affects the number of cTfr cells in kidney transplant recipients. The observed profound, long lasting effects by these agents might dysregulate cTfr functions.