Achieving Donor-Specific Hyporesponsiveness Is Associated with FOXP3+ Regulatory T Cell Recruitment in Human Renal Allograft Infiltrates

Abstract

Exploring new immunosuppressive strategies inducing donor-specific hyporesponsiveness is an important challenge in transplantation. For this purpose, a careful immune monitoring and graft histology assessment is mandatory. Here, we report the results of a pilot study conducted in twenty renal transplant recipients, analyzing the immunomodulatory effects of a protocol based on induction therapy with rabbit anti-thymocyte globulin low doses, sirolimus, and mofetil mycophenolate. Evolution of donor-specific cellular and humoral alloimmune response, peripheral blood lymphocyte subsets and apoptosis was evaluated. Six-month protocol biopsies were performed to assess histological lesions and presence of FOXP3+ regulatory T cells (Tregs) in interstitial infiltrates. After transplantation, there was an early and transient apoptotic effect, mainly within the CD8+HLADR+ T cells, combined with a sustained enhancement of CD4+CD25+high lymphocytes in peripheral blood. The incidence of acute rejection was 35%, all steroid sensitive. Importantly, only pretransplant donor-specific cellular alloreactivity could discriminate patients at risk to develop acute rejection. Two thirds of the patients became donor-specific hyporesponders at 6 and 24 mo, and the achievement of this immunologic state was not abrogated by prior acute rejection episodes. Remarkably, donor-specific hyporesponders had the better renal function and less chronic renal damage. Donor-specific hyporesponsiveness was inhibited by depleting CD4+CD25+high T cells, which showed donor-Ag specificity. FOXP3+CD4+CD25+high Tregs both in peripheral blood and in renal infiltrates were higher in donor-specific hyporesponders than in nonhyporesponders, suggesting that the recruitment of Tregs in the allograft plays an important role for renal acceptance. In conclusion, reaching donor-specific hyporesponsiveness is feasible after renal transplantation and associated with Treg recruitment in the graft.

Presence of FoxP3+ Regulatory T Cells Predicts Outcome of Subclinical Rejection of Renal Allografts

“…17 The number of positive FoxP3 ϩ and CD3 ϩ T cells in the cortex, perivascular areas, and corticomedullary junction aggregates were first counted by using symmetric square power fields (ϫ400; Leica Geosystems, Barcelona, Spain). Then, the proportion of FoxP3 ϩ Treg among the CD3 ϩ T cells (FoxP3 ϩ /CD3 ϩ ) per field was also scored.…”

“…Also, double immunofluorescence labeling was done (Leica TCS-SL), combining the intranuclear FoxP3 with four different T cell surface markers: CD25, CD4, CD8, and CD3 (Master Diagnostica, Vision Biosystem Novocastra), as described previously. 17 All of the histologic analysis was done in a blinded manner.…”

Transient Low-Dose Methotrexate Induces Tolerance to Murine Anti-Thymocyte Globulin and Together They Promote Long-Term Allograft Survival

“…In fact, although T regulatory cells associated with transplantation tolerance (24,30,31,43) are induced by Thymoglobulin and mATG (23,25,26,32,33) and are responsible for mATGmediated prolonged graft survival (16), we did not see increases in T regulatory cells with the mATG and methotrexate combination over that of mATG alone. In addition, there was no evidence of enhanced depletion of T cells, and only a modest extension of mATG effects was observed as a result of blockade of anti-drug Ab responses by methotrexate.…”

“…In particular, as T regulatory cells have been associated with longterm graft survival in transplantation (23,24,30,31), are induced by Thymoglobulin and mATG (22,25,26,32,33), and have previously been demonstrated to be responsible for delayed graft rejection following mATG (16), CD3 + Foxp3 + cells, which bear a phenotype consistent with regulatory T cells, were evaluated. To assess histologic changes in long-surviving grafts, cardiac grafts were collected from the mATG and methotrexate combinationtreated group and the untreated syngeneic group at least 100 d after transplantation.…”

Mechanisms of Murine Spontaneous Liver Transplant Tolerance

“…The immunosuppressive function of Treg cells may occur by cell-cell contact, mediated by the costimulatory molecule cytotoxic T lymphocyte antigen 4, and programmed death ligand 1, and by soluble immunosuppressive factors such as IL-10 and TGF-β. 65,66 The Treg cells can directly suppress the effector CD4+ and CD8+ T cells by reducing IL-2 production and inducing activated T-cell apoptosis. 67 The cytokine IL-2, a growth factor of T cells, is important for Treg cell development and survival.…”