Relative Resistance of Human CD4+ Memory T Cells to Suppression by CD4+CD25+ Regulatory T Cells

Successful expansion of functional CD4⁺CD25⁺ regulatory T cells (T_reg) ex vivo under good manufacturing practice conditions has made T_reg‐cell therapy in clinical transplant tolerance induction a feasible possibility. In animals, T_reg cells home to both transplanted tissues and local lymph nodes and are optimally suppressive if active at both sites. Therefore, they have the opportunity to suppress both naïve and memory CD4⁺CD25^? T cells (Tresp). Clinical transplantation commonly involves depleting therapy at induction (e.g. anti‐CD25), which favors homeostatic expansion of memory T cells. Animal models suggest that T_reg cells are less suppressive on memory, compared with naïve Tresp that mediate allograft rejection. As a result, in the context of human T_reg‐cell therapy, it is important to define the effectiveness of T_reg cells in regulating naïve and memory Tresp. Therefore, we compared suppression of peripheral blood naïve and memory Tresp by fresh and ex vivo expanded T_reg cells using proliferation, cytokine production and activation marker expression (CD154) as readouts. With all readouts, naïve human Tresp were more suppressible by approximately 30% than their memory counterparts. This suggests that T_reg cells may be more efficacious if administered before or at the time of transplantation and that depleting therapy should be avoided in clinical trials of T_reg cells.     

The Potency of Allospecific Tregs Cells Appears to Correlate With T Cell Receptor Functional Avidity

CD4⁺CD25⁺ regulatory T cells (T_reg cells) are an attractive adoptive cell therapy in mediating transplantation tolerance. T‐cell receptor (TcR) activation is critical for T_reg function, suggesting that the TcR avidity of T_reg cells used in therapy may affect the therapeutic outcome. To address this, we compared the regulatory capacity of T_reg lines expressing TcRs derived from two TcR transgenic mice shown to have the same specificity but different functional avidities. T_reg lines generated from CD4⁺CD25⁺ T cells from C57BL/6 mice were transduced with one of either of these TcRs. The antigen specificity of the transduced T_reg lines was confirmed in vitro. T_reg lines expressing the TcR with higher functional avidity showed stronger suppressive capacity in a linked suppression model in vitro. Furthermore, the same T_reg lines demonstrated a stronger proliferation in vivo following antigen exposure. Pretreatment of recipient BL/6 mice with these T_reg cells, together with anti‐CD8 antibody and Rapamycin therapies, prolonged survival of BALB/c skins, as compared with mice that received T_reg lines with lower TcR avidity. Taken together, these data suggest that the TcR functional avidity may be important for T_reg function. It highlights the fact that strategies to select T_reg with higher functional avidity might be beneficial for immunotherapy in transplantation.     

L‐Selectin Is Dispensable for T Regulatory Cell Function Postallogeneic Bone Marrow Transplantation

In murine models, the adoptive transfer of CD4⁺/CD25⁺ regulatory T cells (T_regs) inhibited graft‐versus‐host disease (GvHD). Previous work has indicated a critical role for the adhesion molecule L‐selectin (CD62L) in the function of T_regs in preventing GvHD. Here we examined the capacity of naive wild‐type (WT), CD62L^?/? and ex vivo expanded CD62L^Lo T_regs to inhibit acute GvHD. Surprisingly, we found that CD62L^?/? T_regs were potent suppressors of GvHD, whereas CD62L^Lo T_regs were unable to inhibit disease despite being functionally competent to suppress allo T cell responses in vitro. Concomitant with improved outcomes, WT and CD62L^?/? T_regs significantly reduced liver pathology and systemic pro‐inflammatory cytokine production, although CD62L^?/? T_regs were less effective in reducing lung pathology. While accumulation of CD62L^?/? T_regs in GvHD target organs was equivalent to WT T_regs, CD62L^?/? T_regs did not migrate as well as WT T_regs to peripheral lymph nodes (PLNs) over the first 2 weeks posttransplantation. This work demonstrated that CD62L was dispensable for T_reg‐mediated protection from GvHD.     

Increased frequency of regulatory T Cells and selection of highly potent CD62L+ cells during treatment of human lung transplant recipients with rapamycin

The currently available immunosuppressive agents applied in human transplantation medicine are highly potent in the protection from acute allograft rejection. However, long‐term allograft survival is still poor as these drugs fail to sufficiently prevent chronic allograft rejection. Naturally occurring regulatory T cells have been postulated as the key players to establish long‐lasting transplantation tolerance. Thus, the development of immunosuppressive regimens which shift the pathological balance of cytopathic versus regulatory T cells of human allograft recipients towards a protective T‐cell composition is a promising approach to overcome limitations of current transplantation medicine. Thirty‐three patients that received rapamycin (RPM) or calcineurin inhibitor treatment following lung transplantation were included and their T‐cell compartments analysed. Twelve healthy volunteers without history of lung disease served as controls. In this article, we show that treatment of human lung transplant recipients with RPM is associated with an increased frequency of regulatory T cells, as compared with treatment with calcineurin inhibitors or to healthy controls. Moreover, regulatory T cells during treatment with RPM were CD62Lhigh, a phenotype that displayed an enhanced immunosuppressive capacity ex vivo. Our data support the use of RPM in human lung transplant recipients and undertaking of further prospective studies evaluating its impact on allograft and patient survival.     

Intragraft Regulatory T Cells in Protocol Biopsies Retain Foxp3 Demethylation and Are Protective Biomarkers for Kidney Graft Outcome

Presence of subclinical rejection (SCR) with IF/TA in protocol biopsies of renal allografts has been shown to be an independent predictor factor of graft loss. Also, intragraft Foxp3+ T_reg cells in patients with SCR has been suggested to differentiate harmful from potentially protective infiltrates. Nonetheless, whether presence of Foxp3 T_reg cells in patients with SCR and IF/TA may potentially protect from a deleterious graft outcome has not yet been evaluated. This is a case‐control study in which 37 patients with the diagnosis of SCR and 68 control patients with no cellular infiltrates at 6‐month protocol biopsies matched for age and time of transplantation were evaluated. We first confirmed that numbers of intragraft Foxp3‐expressing T cells in patients with SCR positively correlates with Foxp3 demethylation at the T_reg‐specific demethylation region. Patients with SCR without Foxp3+ T_reg cells within graft infiltrates showed significantly worse 5‐year graft function evolution than patients with SCR and Foxp3+ T_reg cells and those without SCR. When presence of SCR and IF/TA were assessed together, presence of Foxp3+ T_reg could discriminate a subgroup of patients showing the same graft outcome as patients with a normal biopsy. Thus, presence of Foxp3+ T_reg cells in patients with SCR even with IF/TA is associated with a favorable long‐term allograft outcome.     

Contrasting Effects of Cyclosporine and Rapamycin in De Novo Generation of Alloantigen-Specific Regulatory T Cells

The outcome of T‐cell‐mediated responses, immunity or tolerance, critically depends on the balance of cytopathic versus regulatory T (T_reg) cells. In the creation of stable tolerance to MHC incompatible allografts, reducing the unusually large mass of donor‐reactive cytopathic T effector (T_eff) cells via apoptosis is often required. Cyclosporine (CsA) blocks activation‐induced cell death (AICD) of T_eff cells, and is detrimental to tolerance induction by costimulation blockade, whereas Rapamycin (RPM) preserves AICD, and augments the potential of costimulation blockade to create tolerance. While differences between CsA and RPM in influencing apoptosis of activated graft‐destructive T_eff cells are apparent, their effects on graft‐protective T_reg cells remain enigmatic. Moreover, it is unclear whether tolerizing regimens foster conversion of naïve peripheral T cells into alloantigen‐specific T_reg cells for graft protection. Here we show, using reporter mice for T_reg marker Foxp3, that RPM promotes de novo conversion of alloantigen‐specific T_reg cells, whereas CsA completely inhibits this process. Upon transfer, in vivo converted T_reg cells potently suppress the rejection of donor but not third party skin grafts. Thus, the differential effects of RPM and CsA on T_eff and T_reg cells favor the use of RPM in shifting the balance of aggressive to protective type alloimmunity.     

Induction of bona fide regulatory T cells after liver transplantation - the potential influence of polyclonal antithymocyte globulin

T‐cell‐depleting strategies are an integral part of immunosuppressive regimens used in the hematological and solid organ transplant setting. Besides prevention of alloreactivity, treatment with rabbit antithymocyte globulin (rATG) has been related to the induction of immunoregulatory T cells (Treg) in vitro and in vivo. To investigate Treg induced by rATG, we prospectively studied the effect of rATG induction therapy in liver‐transplanted recipients in vivo (n = 28). Treg induction was further evaluated by means of Treg‐specific demethylation region (TSDR) analysis within the FOXP3 locus. Whereas no induction of CD4⁺ CD25^highCD127^? Treg could be observed by phenotypic analysis, we could demonstrate an induction of TSDR⁺ T cells within CD4⁺ T cells exclusively for rATG‐treated patients in the long‐term (day 540) compared with controls (P = NS). Moreover, although in vitro experiments confirm that rATG primarily led to a conversion of CD4⁺ CD25^? into CD4⁺ CD25⁺ T cells displaying immunosuppressive capacities, these cells cannot be classified as bona fide Treg based on their FOXP3 demethylation pattern. Consequently, the generation of Treg after rATG co‐incubation in vitro does not reflect the mechanisms of Treg induction in vivo and therefore the potential clinical relevance of these cells for transplant outcome remains to be determined.     

Belatacept‐Resistant Rejection Is Associated With CD28+ Memory CD8 T Cells

Recently, newer therapies have been designed to more specifically target rejection in an effort to improve efficacy and limit unwanted toxicity. Belatacept, a CD28‐CD80/86 specific reagent, is associated with superior patient survival and graft function compared with traditional therapy, but its adoption as a mainstay immunosuppressive therapy has been tempered by increased rejection rates. It is essential that the underlying mechanisms associated with this rejection be elucidated before belatacept is more widely used. To that end, we designed a study in a nonhuman primate kidney transplant model where animals were treated with either a belatacept‐ or a tacrolimus‐based immunosuppressive regimen. Interestingly, we found that elevated pretransplant frequencies of CD28⁺CD8⁺T_EMRA cells are associated with rejection on belatacept but not tacrolimus treatment. Further analysis showed that the CD28⁺CD8⁺T_EMRA cells rapidly lose CD28 expression after transplant in those animals that go on to reject with the allograft infiltrate being predominantly CD28^?. These data suggest that CD28⁺ memory T cells may be resistant to belatacept, capable of further differentiation including loss of CD28 expression while maintaining effector function. The unique signaling requirements of CD28⁺ memory T cells provide opportunities for the development of targeted therapies, which may synergize with belatacept to prevent costimulation‐independent rejection.     

Depletion of CD4+CD25+ Regulatory T Cells Promotes a Tumor-Specific Immune Response in Pancreas Cancer–Bearing Mice

Background Pancreas cancer–bearing mice have an increased prevalence of immunosuppressive CD4⁺CD25⁺ regulatory T cells (T_reg). Depletion of T_reg results in smaller tumors and prolonged host survival. The objective of this study was to evaluate the tumor-specific immune response after depletion of T_reg alone or in combination with a cancer vaccine.Methods Four groups of C57BL/6 mice were challenged with pancreas adenocarcinoma cells (Pan02). The mice received four combinations of antibody-mediated T_reg depletion and whole tumor cell vaccination: (1) no treatment, (2) T_reg depletion only, (3) vaccination only, or (4) T_reg depletion and vaccination. Splenocytes and lymphocytes from tumor-draining lymph nodes were analyzed for tumor-specific release of interferon γ by enzyme-linked immunosorbent spot assay.Results In T_reg-depleted and vaccinated mice, a strong statistical trend toward smaller tumors (P = .05) and longer survival (P = .054) was found compared with untreated mice. T_reg-depleted mice showed significantly more tumor-specific cells than undepleted mice (P = .02). The number of tumor-specific cells was significantly higher in tumor-draining lymph nodes than in the spleen (P = .002). Similarly, significantly more tumor-specific cells were found in spleens of T_reg-depleted and vaccinated mice than in vaccinated-only mice (P = .009).Conclusions Depletion of T_reg alone or in combination with a whole tumor cell vaccine promotes a tumor-specific immune response. Thus, strategies incorporating T_reg depletion might improve the efficacy of cancer vaccines.Presented at the 57th Annual Cancer Symposium of the Society of Surgical Oncology, New York, New York, March 18–21, 2004.     

Glutamate Receptor Interacting Protein 1 Regulates CD4+ CTLA‐4 Expression and Transplant Rejection

PDZ domains are common 80‐ to 90‐amino‐acid regions named after the first three proteins discovered to share these domains: postsynaptic density 95, discs large, and zonula occludens. PDZ domain‐containing proteins typically interact with the C‐terminus of membrane receptors. Glutamate receptor interacting protein 1 (GRIP1), a seven–PDZ domain protein scaffold, regulates glutamate receptor surface expression and trafficking in neurons. We have found that human and mouse T cells also express GRIP1. T cell–specific GRIP1^?/? mice >11 weeks old had prolonged cardiac allograft survival. Compared with wild‐type T cells, in vitro stimulated GRIP1^?/? T cells had decreased expression of activation markers and increased apoptotic surface marker expression. Surface expression of the strong T cell inhibitory molecule cytotoxic T lymphocyte antigen‐4 (CTLA‐4) was increased on GRIP1^?/? T cells from mice >11 weeks old. CTLA‐4 increases with T cell stimulation and its surface expression on GRIP1^?/? T cells remained high after stimulation was removed, indicating a possible internalization defect in GRIP1‐deficient T cells. CTLA‐4–blocking antibody treatment following heart transplantation led to complete rejection in T cell GRIP1^?/? mice, indicating that increased CTLA‐4 surface expression contributed to the extended graft survival. Our data indicate that GRIP1 regulates T cell activation by regulating CTLA‐4 surface expression.     

B cells assist allograft rejection in the deficiency of protein kinase c‐theta

We have previously shown that mice deficient in protein kinase C theta (PKCθ) have the ability to reject cardiac allografts, but are susceptible to tolerance induction. Here we tested role of B cells in assisting alloimmune responses in the absence of PKCθ. Mouse cardiac allograft transplantations were performed from Balb/c (H‐2d) to PKCθ knockout (PKCθ^?/?), PKCθ and B cell double‐knockout (PBDK, H‐2b) mice and wild‐type (WT) C57BL/6 (H‐2b) mice. PBDK mice spontaneously accepted the allografts with the inhibition of NF‐κB activation in the donor cardiac allograft. Anti‐B cell antibody (rituximab) significantly delayed allograft rejection in PKCθ^?/?, but not in WT mice. Co‐transfer of PKCθ^?/? T plus PKCθ^?/? B cells or primed sera triggered allograft rejection in Rag1^?/? mice, and only major histocompatibility complex class II‐enriched B cells, but not class I‐enriched B cells, were able to promote rejection. This, together with the inability of PKCθ^?/? and CD28^?/? double‐deficient (PCDK) mice to acutely reject allografts, suggested that an effective cognate interaction between PKCθ^?/? T and B cells for acute rejection is CD28 molecule dependent. We conclude that T–B cell interactions synergize with PKCθ^?/? T cells to mediate acute allograft rejection.     

Dominant regulation of long‐term allograft survival is mediated by microRNA‐142

Organ transplantation is often lifesaving, but the long‐term deleterious effects of combinatorial immunosuppression regimens and allograft failure cause significant morbidity and mortality. Long‐term graft survival in the absence of continuing immunosuppression, defined as operational tolerance, has never been described in the context of multiple major histocompatibility complex (MHC) mismatches. Here, we show that miR‐142 deficiency leads to indefinite allograft survival in a fully MHC mismatched murine cardiac transplant model in the absence of exogenous immunosuppression. We demonstrate that the cause of indefinite allograft survival in the absence of miR‐142 maps specifically to the T cell compartment. Of therapeutic relevance, temporal deletion of miR‐142 in adult mice prior to transplantation of a fully MHC mismatched skin allograft resulted in prolonged allograft survival. Mechanistically, miR‐142 directly targets Tgfbr1 for repression in regulatory T cells (T_REG). This leads to increased T_REG sensitivity to transforming growth factor – beta and promotes transplant tolerance via an augmented peripheral T_REG response in the absence of miR‐142. These data identify manipulation of miR‐142 as a promising approach for the induction of tolerance in human transplantation.     

Intragraft CD11b+IDO+ Cells Mediate Cardiac Allograft Tolerance by ECDI‐Fixed Donor Splenocyte Infusions

We have previously shown that pre‐ and post‐transplant infusions of donor splenocytes treated with 1‐ethyl‐3‐(3’‐dimethylaminopropyl)‐carbodiimide (ECDI‐SPs) provide permanent donor‐specific protection of islet allografts. The efficacy of donor ECDI‐SPs in protecting vascularized cardiac allografts and mechanism(s) of protection are unknown. In this study, we show that infusions of ECDI‐SPs significantly prolong cardiac allograft survival concomitant with an impressive accumulation of CD11b⁺IDO⁺ cells in the cardiac allograft, and that the presence of this population is dependent on Gr1⁺ cells. Consequently, depletion of Gr1⁺ cells or inhibition of indoleamine 2,3 dioxygenase (IDO) activity abrogates graft protection by ECDI‐SPs infusions. In addition, T cells from ECDI‐SPs treated recipients secrete high levels of interleukin 10 and interleukin 13 upon in vitro restimulation, which are also dampened in recipients treated with the IDO inhibitor. Furthermore, combination of donor ECDI‐SPs with a short course of rapamycin provides indefinite cardiac allograft survival in 100% of the recipients. These findings reveal a novel mechanism of donor ECDI‐SPs in inducing cardiac transplant tolerance and provide several targets that are amenable to therapeutic manipulations for tolerance induction for cardiac transplantation.     

Increased Pretransplant Frequency of CD28+ CD4+ TEM Predicts Belatacept‐Resistant Rejection in Human Renal Transplant Recipients

While most human T cells express the CD28 costimulatory molecule constitutively, it is well known that age, inflammation, and viral infection can drive the generation of CD28^null T cells. In vitro studies have demonstrated that CD28^null cell effector function is not impacted by the presence of the CD28 costimulation blocker belatacept. As such, a prevailing hypothesis suggests that CD28^null cells may precipitate costimulation blockade‐resistant rejection. However, CD28⁺ cells possess more proliferative and multifunctional capacity, factors that may increase their ability to successfully mediate rejection. Here, we performed a retrospective immunophenotypic analysis of adult renal transplant recipients who experienced acute rejection on belatacept treatment as compared to those who did not. Intriguingly, our findings suggest that patients possessing higher frequency of CD28⁺ CD4⁺ T_EM prior to transplant were more likely to experience acute rejection following treatment with a belatacept‐based immunosuppressive regimen. Mechanistically, CD28⁺ CD4⁺ T_EM contained significantly more IL‐2 producers. In contrast, CD28^null CD4⁺ T_EM isolated from stable belatacept‐treated patients exhibited higher expression of the 2B4 coinhibitory molecule as compared to those isolated from patients who rejected. These data raise the possibility that pretransplant frequencies of CD28⁺ CD4⁺ T_EM could be used as a biomarker to predict risk of rejection following treatment with belatacept.     

Prolongation of Cardiac Allograft Survival by a Novel Population of Autologous CD117+ Bone Marrow‐Derived Progenitor Cells

Autologous CD117⁺ progenitor cells (PC) have been successfully utilized in myocardial infarction and ischemic injury, potentially through the replacement/repair of damaged vascular endothelium. To date, such cells have not been used to enhance solid organ transplant outcome. In this study, we determined whether autologous bone marrow‐derived CD117⁺PC could benefit cardiac allograft survival, possibly by replacing donor vascular cells. Autologous, positively selected CD117⁺PC were administered posttransplantation and allografts were assessed for acute rejection. Although significant generation of recipient vascular cell chimerism was not observed, transferred PC disseminated both to the allograft and to peripheral lymphoid tissues and facilitated a significant, dose‐dependent prolongation of allograft survival. While CD117⁺PC dramatically inhibited alloreactive T cell proliferation in vitro, this property did not differ from nonprotective CD117^? bone marrow populations. In vivo, CD117⁺ PC did not significantly inhibit T cell alloreactivity or increase peripheral regulatory T cell numbers. Thus, rather than inhibiting adaptive immunity to the allograft, CD117⁺ PC may play a cytoprotective role in prolonging graft survival. Importantly, autologous CD117⁺PC appear to be distinct from bone marrow‐derived mesenchymal stem cells (MSC) previously used to prolong allograft survival. As such, autologous CD117⁺PC represent a novel cellular therapy for promoting allograft survival.     

RIPK3‐Mediated Necroptosis Promotes Donor Kidney Inflammatory Injury and Reduces Allograft Survival

Kidney transplant injury occurs with ischemia and alloimmunity. Members of the receptor interacting protein kinase family (RIPK1,3) are key regulators of “necroptosis,” a newly recognized, regulated form of necrosis. Necroptosis and apoptosis death appear to be counterbalanced as caspase‐8 inhibition can divert death from apoptosis to necrosis. Inhibition of necroptosis in donor organs to limit injury has not been studied in transplant models. In this study, necroptosis was triggered in caspase inhibited tubular epithelial cells (TEC) exposed to tumor necrosis factor alpha in vitro, while RIPK1 inhibition with necrostatin‐1 or use of RIPK3^?/? TEC, prevented necroptosis. In vivo, short hairpin RNA silencing of caspase‐8 in donor B6 mouse kidneys increased necroptosis, enhanced high‐mobility group box 1 release, reduced renal function and accelerated rejection when transplanted into BALB/c recipients. Using ethidium homodimer perfusion to assess necrosis in vivo, necrosis was abrogated in RIPK3^?/? kidneys postischemia. Following transplantation, recipients receiving RIPK3^?/? kidneys had longer survival (p = 0.002) and improved renal function (p = 0.03) when compared to controls. In summary, we show for the first time that RIPK3‐mediated necroptosis in donor kidneys can promote inflammatory injury, and has a major impact on renal ischemia–reperfusion injury and transplant survival. We suggest inhibition of necroptosis in donor organs may similarly provide a major clinical benefit.

     

Loss of DRAK2 Signaling Enhances Allogeneic Transplant Survival by Limiting Effector and Memory T Cell Responses

Here, we demonstrate that loss of DRAK2 signaling significantly promotes the acceptance of allogeneic engraftment in two separate transplant models without promoting generalized immunosuppression. Drak2?/? T cells failed to reject allogeneic tumors, and were defective in rejecting Balb/C allogeneic skin grafts on C57BL6/J recipients. A significant fraction of alloreactive Drak2?/? T cells underwent apoptosis following activation, whereas enforced expression of Bcl‐xL in Drak2?/? T cells restored allograft rejection. Formation of allogeneic memory was also greatly hampered in T cells lacking the Drak2 gene. Adoptive transfer of memory T cells from Drak2?/? mice failed to promote the rejection of allogeneic tumors, and such cells led to significantly delayed rejection of skin allografts in the Balb/C‐>C57BL/6J model. Costimulatory blockade by in vivo administration of Cytotoxic T‐Lymphocyte Antigen 4 fusion protein (CTLA4‐Ig) synergized with the DRAK2 deficiency and led to long‐term allogeneic skin graft acceptance. Overall, these results demonstrate that DRAK2 plays an important role in primary and memory T cell responsiveness to allografts. Because previous studies have demonstrated that a loss of DRAK2 does not negatively impact antiviral immunity, the studies here underscore the potential utility of pharmacological blockade of DRAK2 to achieve transplant maintenance without the imposition of generalized immunosuppression.     

In Vivo Development of Transplant Arteriosclerosis in Humanized Mice Reflects Alloantigen Recognition and Peripheral Treg Phenotype of Lung Transplant Recipients

Experimentally, regulatory T cells inhibit rejection. In clinical transplantations, however, it is not known whether T cell regulation is the cause for, or an epiphenomenon of, long‐term allograft survival. Here, we study naïve and alloantigen‐primed T cell responses of clinical lung transplant recipients in humanized mice. The pericardiophrenic artery procured from human lung grafts was implanted into the aorta of NODrag^?/?/IL‐2rγc^?/? mice reconstituted with peripheral blood mononuclear cells (PBMCs) from the respective lung recipient. Naïve or primed allogeneic PBMCs procured 21 days post–lung transplantation with or without enriching for CD4⁺CD25^high T cells were used. Transplant arteriosclerosis was assessed 28 days later by histology. Mice reconstituted with alloantigen‐primed PBMCs showed significantly more severe transplant arteriosclerosis than did mice with naïve PBMCs (p = 0.005). Transplant arteriosclerosis was equally suppressed by enriching for autologous naïve (p = 0.012) or alloantigen‐primed regulatory T cells (Tregs) (p = 0.009). Alloantigen priming in clinical lung recipients can be adoptively transferred into a humanized mouse model. Transplant arteriosclerosis elicited by naïve or alloantigen‐primed PBMCs can be similarly controlled by potent autologous Tregs. Cellular therapy with expanded autologous Tregs in lung transplantation might be a promising future strategy.