An ultimate goal in transplantation is induction of immune tolerance to transplants, a state of unresponsiveness to the allograft in a competent immune system without immunosuppression. Albeit rare, operational tolerance, generally defined as stable graft function without clinical features of chronic rejection in the absence of any immunosuppressive drugs for at least 1 year, has been observed in kidney transplant recipients after intentional weaning of immunosuppression or in nonadherent patients who have stopped their medication.1 Whereas some of these patients have been off immunosuppression for more than 5 years without signs of rejection, a number of patients developed rejection at some point after withdrawal, which in most cases resulted in loss of the graft. Unraveling immunologic mechanisms that play a role in the development of operational tolerance and defining biomarkers and predictors of tolerance is paramount for the development of strategies to achieve operational tolerance in a safe manner.
Various studies have investigated the transcriptional profile and/or immunophenotypic characteristics of operational tolerant kidney transplant recipients in comparison to healthy individuals, kidney transplant patients with chronic rejection and/or patients with stable function on immunosuppression. Unique peripheral blood mononuclear cell gene signatures associated with operational tolerance in kidney transplant patients have been described.2-4 Differences in the B-cell compartment were observed in tolerant kidney transplant recipients compared with the other groups.3,4 Frequencies of activated CD4+ T cells as well as gene expression levels of markers associated with T-cell activation and proinflammatory Th1/Th2 responses were lower in tolerant patients than in chronically rejecting patients.2,3 Additionally, frequencies of CD25highCD4+ T cells and FOXP3 expression levels in tolerant patients were comparable to that in healthy individuals, whereas patients with chronic rejection had significantly lower levels.3,5 Whether these alterations in B- and T-cell compartment play a role in the induction of tolerance in operational tolerant patients, or whether they contribute to the maintenance of tolerance in the absence of immunosuppression, remains to be elucidated.
In this issue of Transplantation, Nova-Lamperti et al6 show that tolerant kidney transplant recipients have an altered peripheral Th17 compartment compared with healthy controls and patients with chronic rejection. Gene expression analysis revealed lower levels of retinoic acid receptor-related orphan receptor C, a master regulator of Th17 cells, and phenotypic analysis showed a lower proportion of Th17 cells in tolerant kidney transplant recipients versus healthy controls and chronically rejecting patients. In addition, the authors describe an impaired Th17 polarization in tolerant recipients which may be attributed to reduced extracellular signal-regulated kinase (ERK) phosphorylation after T-cell stimulation. The ERK pathway, one of the major signaling cassettes of the mitogen-activated protein kinase (MAPK) signaling pathway, is important for various cellular processes, including cell proliferation and differentiation. Interestingly, Nova-Lamperti et al7 recently demonstrated reduced ERK phosphorylation in B cells of tolerant patients after B-cell stimulation as well. These findings suggest that an impaired ERK/MAPK pathway may be important for tolerance induction.
This study sheds new light on immunologic pathways that may play a role in the development and/or maintenance of operational tolerance. The findings give rise to several crucial questions and create a “chicken-or-the-egg” dilemma. Did the altered peripheral Th17 compartment contribute to the induction of tolerance in these patients or is it merely the result of other immunoregulatory mechanisms that were key in the tolerance induction? Did it develop after transplantation, possibly due to the effects of immunosuppression or graft-host interaction, or was it “naturally” occurring and were these patients therefore more prone to tolerance induction? It would be interesting to investigate peripheral blood samples of the tolerant patient group at earlier time points to assess the Th17 compartment before tolerance induction or even before transplantation. Viklicky et al8 studied prospectively several genes associated with tolerance in kidney transplant recipients on immunosuppression and observed an upregulation of B cell-related genes in rejection-free patients, as seen in tolerant patients, compared with patients undergoing rejection. In addition, Heidt et al9 demonstrated that B cell-related genes associated with operational tolerance are downregulated during an episode of acute rejection. Because the gene expression levels of these B cell-related biomarkers seemed to be comparable before transplantation, these findings suggest that the observed differences between rejection-free and rejecting patients may develop after transplantation.
Another question is whether the findings of Nova-Lamperti et al reflect donor-specific unresponsiveness or if it is a broad immune deficiency? The setup of the experiments suggests the latter as a generic stimulation was used for activation and differentiation. The authors describe that no evidence of increased infections or cancer have been found for this population, but further analysis is warranted. Ballet et al10 found a heterogeneous response of tolerant kidney transplant recipients to influenza vaccination; some tolerant patients presented humoral and cellular responses that were comparable to those in healthy controls, whereas others had poor humoral responses comparable to immunosuppressed patients. Albeit in a small study group, this study demonstrated that a certain degree of global immune deficiency may be present in (at least) some tolerant patients.
The difficulty of comparison studies is the clinical situation of the different groups: healthy controls share the absence of immunosuppression with tolerant patients but have not received a transplant and presumably do not have a history of immunosuppressive treatment; chronically rejecting patients have received a transplant but also receive immunosuppressive medication and may have an activated immune system. A study by Rebollo-Mesa et al11 demonstrated that immunosuppressive drugs can bias the expression of genes and affect the frequency of cell subsets in peripheral blood, indicating that immunosuppression can be a confounding factor. Although essential, comparison studies may not directly identify immunologic mechanisms that contribute to the development and maintenance of operational tolerance. Longitudinal studies (preferably with a prospective approach) will be required to assess whether alterations in B- and T-cell compartments are “the chicken or the egg” in the complex phenomenon of operational tolerance.
1. Orlando G, Hematti P, Stratta RJ, et al. Clinical operational tolerance after renal transplantation: current status and future challenges. Ann Surg
2. Brouard S, Mansfield E, Braud C, et al. Identification of a peripheral blood transcriptional biomarker panel associated with operational renal allograft tolerance. Proc Natl Acad Sci U S A
3. Sagoo P, Perucha E, Sawitzki B, et al. Development of a cross-platform biomarker signature to detect renal transplant tolerance in humans. J Clin Invest
4. Newell KA, Asare A, Kirk AD, et al. Identification of a B cell signature associated with renal transplant tolerance in humans. J Clin Invest
5. Louis S, Braudeau C, Giral M, et al. Contrasting CD25hiCD4 + T cells/FOXP3 patterns in chronic rejection and operational drug-free tolerance. Transplantation
6. Nova-Lamperti E, Romano M, Christakoudi S, et al. Reduced TCR-signalling contributes to impaired Th17 responses in tolerant kidney transplant recipients. Transplantation
7. Nova-Lamperti E, Chana P, Mobillo P, et al. Increased CD40 ligation and reduced BCR signalling leads to higher IL-10 production in B cells from tolerant kidney transplant patients. Transplantation
8. Viklicky O, Krystufkova E, Brabcova I, et al. B-cell-related biomarkers of tolerance are up-regulated in rejection-free kidney transplant recipients. Transplantation
9. Heidt S, Vergunst M, Anholts JD, et al. B cell markers of operational tolerance can discriminate acute kidney allograft rejection from stable graft function. Transplantation
10. Ballet C, Roussey-Kesler G, Aubin JT, et al. Humoral and cellular responses to influenza vaccination in human recipients naturally tolerant to a kidney allograft. Am J Transplant
11. Rebollo-Mesa I, Nova-Lamperti E, Mobillo P, et al. Biomarkers of tolerance in kidney transplantation: are we predicting tolerance or response to immunosuppressive treatment? Am J Transplant