Blood CD9+ B cell,a biomarker of bronchiolitis obliterans syndrome after lung transplantation

Bronchiolitis obliterans syndrome is the main limitation for long‐term survival after lung transplantation. Some specific B cell populations are associated with long‐term graft acceptance. We aimed to monitor the B cell profile during early development of bronchiolitis obliterans syndrome after lung transplantation. The B cell longitudinal profile was analyzed in peripheral blood mononuclear cells from patients with bronchiolitis obliterans syndrome and patients who remained stable over 3 years of follow‐up. CD24^hiCD38^hi transitional B cells were increased in stable patients only, and reached a peak 24 months after transplantation, whereas they remained unchanged in patients who developed a bronchiolitis obliterans syndrome. These CD24^hiCD38^hi transitional B cells specifically secrete IL‐10 and express CD9. Thus, patients with a total CD9⁺ B cell frequency below 6.6% displayed significantly higher incidence of bronchiolitis obliterans syndrome (AUC = 0.836, PPV = 0.75, NPV = 1). These data are the first to associate IL‐10‐secreting CD24^hiCD38^hi transitional B cells expressing CD9 with better allograft outcome in lung transplant recipients. CD9‐expressing B cells appear as a contributor to a favorable environment essential for the maintenance of long‐term stable graft function and as a new predictive biomarker of bronchiolitis obliterans syndrome–free survival.     

A Pilot Trial Targeting the ICOS–ICOS‐L Pathway in Nonhuman Primate Kidney Transplantation

Costimulation blockade with the B7‐CD28 pathway‐specific agent belatacept is now used in clinical kidney transplantation, but its efficacy remains imperfect. Numerous alternate costimulatory pathways have been proposed as targets to synergize with belatacept, one of which being the inducible costimulator (ICOS)–ICOS ligand (ICOS‐L) pathway. Combined ICOS–ICOS‐L and CD28‐B7 blockade has been shown to prevent rejection in mice, but has not been studied in primates. We therefore tested a novel ICOS‐Ig human Fc‐fusion protein in a nonhuman primate (NHP) kidney transplant model alone and in combination with belatacept. ICOS‐Ig did not prolong rejection‐free survival as a monotherapy or in combination with belatacept. In ICOS‐Ig alone treated animals, most graft‐infiltrating CD4⁺ and CD8⁺ T cells expressed ICOS, and ICOS⁺ T cells were present in peripheral blood to a lesser degree. Adding belatacept reduced the proportion of graft‐infiltrating ICOS⁺ T cells and virtually eliminated their presence in peripheral blood. Graft‐infiltrating T cells in belatacept‐resistant rejection were primarily CD8⁺CD28^?, but importantly, very few CD8⁺CD28^? T cells expressed ICOS. We conclude that ICOS‐Ig, alone or combined with belatacept, does not prolong renal allograft survival in NHPs. This may relate to selective loss of ICOS with CD28 loss.

     

B cell reconstitution following alemtuzumab induction under a belatacept‐based maintenance regimen

Lymphocyte depletion has been shown to control costimulation blockade‐resistant rejection but, in some settings, to exacerbate antibody‐mediated rejection (AMR). We have used alemtuzumab, which depletes T and B cells, combined with belatacept and rapamycin and previously reported control of both costimulation blockade‐resistant rejection and AMR. To evaluate this regimen's effect on B cell signatures, we investigated 40 patients undergoing this therapy. B cell counts and phenotypes were interrogated using flow cytometry, and serum was analyzed for total IgG, IgM, and donor‐specific alloantibody (DSA). Alemtuzumab induction produced pan‐lymphocyte depletion; B cells repopulated faster and more completely than T cells. Reconstituting B cells were predominantly naïve, and memory B cells were significantly reduced (P = .001) post repopulation. Two B cell populations with potential immunomodulatory effects—regulatory (CD38^hiCD24^hiIgM^hiCD20^hi) and transitional B cells (CD19⁺CD27^?IgD⁺CD38^hi)—were enriched posttransplant (P = .001). Total serum IgG decreased from baseline (P = .016) while IgM levels remained stable. Five patients developed DSAs within 36 months posttransplant, but none developed AMR. Baseline IgG levels in these patients were significantly higher than those in patients without DSAs. These findings suggest that belatacept and rapamycin together limit homeostatic B cell activation following B cell depletion and may lessen the risk of AMR. This regimen warrants prospective, comparative study. ClinicalTrials.gov NCT00565773.     

Dynamic changes of B‐cell compartments in kidney transplantation: lack of transitional B cells is associated with allograft rejection

B cells play an important role in the immune responses which affect the outcomes of kidney allografts. Dynamic changes of B‐cell compartments in clinical kidney transplantation are still poorly understood. B‐cell subsets were prospectively monitored using flow cytometry for 1 year in 98 kidney transplant recipients. Data were correlated with immunosuppression and clinical outcomes. An increase in the total population of B lymphocytes was observed during the first week after transplantation. The level of IgM^highCD38^highCD24^high transitional B cells reduced significantly up until the third month, with partial repopulation in the first year. Lower numbers of transitional B cells in the third month were associated with higher risk of graft rejection. IgM⁺IgD⁺CD27^? naive B cells did not change within follow‐up. IgM⁺CD27⁺ nonswitched memory B cells and IgM^?CD27⁺ switched memory B cells increased on post‐operative day 7. IgM^?CD38^highCD27^high plasmablasts showed similar kinetics during the first post‐transplant year, similar to transitional B cells. In conclusion, sensitized kidney transplant recipients as well as those with either acute or chronic rejection within the first post‐transplant year exhibited lower levels of transitional B cells. Therefore, these data further support the hypothesis that transitional B cells have a protective role in kidney transplantation.     

Calcineurin and mTOR inhibitors have opposing effects on regulatory T cells while reducing regulatory B cell populations in kidney transplant recipients

BackgroundRegulatory B (Breg) and T (Treg) cells represent a biomarker for tolerance in transplant patients. Despite the importance of Treg and Breg in transplantation and the suggested crosstalk between both suppressive cell populations, little is known on how they are influenced by long-term immunosuppressive treatment. The aim of the present study was to investigate the effect of different immunosuppressive drugs used in routine clinical practice on Treg and Breg cell numbers.MethodsThirty-six kidney transplant recipients with stable graft function were recruited and classified according to their concomitant therapy: 22 patients received calcineurin inhibitors (CNI) and 14 patients received mammalian target of rapamycin (mTOR) inhibitors. A group of 8 healthy untreated subjects was included as control. Absolute numbers of peripheral blood-derived IL10-producing B cells (CD19⁺IL10⁺), CD19⁺CD24^hiCD38^hi transitional B cells and Treg cells (CD4⁺CD25⁺FOXP3⁺) were quantified in all KT patients and controls by flow cytometry.ResultsCD19⁺CD24^hiCD38^hi transitional B cells increased over time and seem to be related with long-term therapeutic graft survival irrespective of the treatment regimen. CNI and mTOR inhibitors significantly reduced numbers of Breg cells when compared with healthy individuals, whereas mTOR inhibitors expanded Treg cells in comparison with CNI drugs.ConclusionsBridging the drug-mediated reduction of Breg cell numbers in vivo with the requirements of Breg cells for long-term transplant success remains an as yet unresolved task for therapeutic intervention. Further larger cohort studies that evaluate the effect of different treatment regimen on defined lymphocyte subpopulations are warranted.     

Immunophenotyping of peripheral immunoregulatory as well as Th17A and Th22 cell subpopulations in kidney transplant recipients under belatacept or cyclosporine treatment

ObjectiveRegulatory Foxp3-expressing T cells (Tregs), IL-10-producing B cells (Bregs), and IDO-expressing dendritic cells (DCregs) downregulate inflammatory processes and induce peripheral tolerance, while Th17A and Th22 cell subpopulations are of proinflammatory nature. The aims of the study were to characterize and to enumerate peripheral Tregs, Bregs, and DCregs and Th17A and Th22 cell subpopulations in kidney transplant recipients (KTRs) under belatacept or cyclosporine treatment.MethodsForty-one KRT patients (30 under belatacept treatment and 11 under cyclosporine treatment) and 26 healthy donors (HDs) were included in the study. CD19⁺-expressing peripheral B lymphocytes were purified by positive selection. IL-10-producing B cells, CD4⁺/CD25^highFoxp3⁺, and CD8⁺/CD28^?Foxp3⁺ Tregs, CCR6⁺/CD123⁺/IDO⁺ DCs, as well as Th17A and Th22 cell subpopulations were quantitated by flow cytometry.ResultsOf the IL-10-producing Bregs, CD19⁺/CD24^high/CD38^high/CD5⁺, CD19⁺/CD24^high/CD38^high/CD10⁺, CD19⁺/CD24^high/CD38^high/CD20⁺, and CD19⁺/CD24^high/CD38^high/CD27^? had significant higher frequency in patients under belatacept treatment when compared with those under cyclosporine. Only CD19⁺/CD24^high/CD38^high/CD27⁺ and CD19⁺/CD24^high/CD38^high/CXCR7⁺ cells had significant higher frequency in patients under cycloporine treatment when compared to those under belatacept. The percentages of IDO-expressing pDC, CD4⁺/CD25^highFoxp3⁺, and CD8⁺/CD28^?Foxp3⁺ were significantly higher in the belatacept group when compared the cyclosporine one, while Th17A and Th22 cells had significant higher frequency in the latter group.ConclusionBelatacept seems to maintain and enhance, at least systemically, a tolerant profile to renal allograft in transplant recipients by means of higher circulatory frequencies of regulatory B, T and pDC subpopulations.     

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.     

Selective Targeting of Human Alloresponsive CD8+ Effector Memory T Cells Based on CD2 Expression

Costimulation blockade (CoB), specifically CD28/B7 inhibition with belatacept, is an emerging clinical replacement for calcineurin inhibitor‐based immunosuppression in allotransplantation. However, there is accumulating evidence that belatacept incompletely controls alloreactive T cells that lose CD28 expression during terminal differentiation. We have recently shown that the CD2‐specific fusion protein alefacept controls costimulation blockade‐resistant allograft rejection in nonhuman primates. Here, we have investigated the relationship between human alloreactive T cells, costimulation blockade sensitivity and CD2 expression to determine whether these findings warrant potential clinical translation. Using polychromatic flow cytometry, we found that CD8⁺ effector memory T cells are distinctly high CD2 and low CD28 expressors. Alloresponsive CD8⁺CD2^hiCD28^? T cells contained the highest proportion of cells with polyfunctional cytokine (IFNγ, TNF and IL‐2) and cytotoxic effector molecule (CD107a and granzyme B) expression capability. Treatment with belatacept in vitro incompletely attenuated allospecific proliferation, but alefacept inhibited belatacept‐resistant proliferation. These results suggest that highly alloreactive effector T cells exert their late stage functions without reliance on ongoing CD28/B7 costimulation. Their high CD2 expression increases their susceptibility to alefacept. These studies combined with in vivo nonhuman primate data provide a rationale for translation of an immunosuppression regimen pairing alefacept and belatacept to human renal transplantation.     

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.     

Immunologic Human Renal Allograft Injury Associates with an Altered IL-10/TNF-α Expression Ratio in Regulatory B Cells

Human B cells with immunoregulatory properties in vitro (Bregs) have been defined by the expression of IL-10 and are enriched in various B-cell subsets. However, proinflammatory cytokine expression in B-cell subsets is largely unexplored. We examined the cytokine profiles of human PBMCs and found that subsets of CD24^hiCD38^hi transitional B cells (TrBs), CD24^hiCD27⁺ memory B cells, and naïve B cells express IL-10 and the proinflammatory cytokine TNF-α simultaneously. TrBs had the highest IL-10/TNF-α ratio and suppressed proinflammatory helper T cell 1 (Th1) cytokine expression by autologous T cells in vitro more potently than memory B cells did, despite similar IL-10 expression. Whereas neutralization of IL-10 significantly inhibited TrB-mediated suppression of autologous Th1 cytokine expression, blocking TNF-α increased the suppressive capacity of both memory and naïve B-cell subsets. Thus, the ratio of IL-10/TNF-α expression, a measure of cytokine polarization, may be a better indicator of regulatory function than IL-10 expression alone. Indeed, compared with TrB cells from patients with stable kidney graft function, TrBs from patients with graft rejection displayed similar IL-10 expression levels but increased TNF-α expression (i.e., reduced IL-10/TNF-α ratio), did not inhibit in vitro expression of Th1 cytokines by T cells, and abnormally suppressed expression of Th2 cytokines. In patients with graft dysfunction, a low IL-10/TNF-α ratio in TrBs associated with poor graft outcomes after 3 years of follow-up. In summary, these results indicate that B cell–mediated immune regulation is best characterized by the cytokine polarization profile, a finding that was confirmed in renal transplant patients.     

TIGIT regulates apoptosis of risky memory T cell subsets implicated in belatacept-resistant rejection

Belatacept confers increased patient and graft survival in renal transplant recipients relative to calcineurin inhibitors, but is associated with an increased rate of acute rejection. Recent immunophenotypic studies comparing pretransplant T cell phenotypes of patients who reject versus those who remain stable on belatacept identified three potential “risky” memory T cell subsets that potentially underlie belatacept-resistant rejection: CD4⁺ CD28⁺ T_EM, CD8⁺ CD28^null, and CD4⁺ CD57⁺ PD1⁻ subsets. Here, we compared key phenotypic and functional aspects of these human memory T cell subsets, with the goal of identifying additional potential targets to modulate them. Results demonstrate that TIGIT, an increasingly well-appreciated immune checkpoint receptor, was expressed on all three risky memory T cell subsets in vitro and in vivo in the presence of belatacept. Coculture of human memory CD4⁺ and CD8⁺ T cells with an agonistic anti-TIGIT mAb significantly increased apoptotic cell death of all three risky memory T cell subsets. Mechanistically, TIGIT-mediated apoptosis of risky memory T cells was dependent on FOXP3⁺ Treg, suggesting that agonism of the TIGIT pathway increases FOXP3⁺ Treg suppression of human memory T cell populations. Overall, these data suggest that TIGIT agonism could represent a new therapeutic target to inhibit belatacept-resistant rejection during transplantation.     

IL‐7 receptor heterogeneity as a mechanism for repertoire change during postdepletional homeostatic proliferation and its relation to costimulation blockade–resistant rejection

Kidney transplant patients treated with belatacept without depletional induction experience higher rates of acute rejection compared to patients treated with conventional immunosuppression. Costimulation blockade–resistant rejection (CoBRR) is associated with terminally differentiated T cells. Alemtuzumab induction and belatacept/sirolimus immunotherapy effectively prevent CoBRR. We hypothesized that cells in late phases of differentiation would be selectively less capable than more naive phenotypes of repopulating postdepletion, providing a potential mechanism by which lymphocyte depletion and repopulation could reduce the risk of CoBRR. Lymphocytes from 20 recipients undergoing alemtuzumab‐induced depletion and belatacept/sirolimus immunosuppression were studied longitudinally for markers of maturation (CCR7, CD45RA, CD57, PD1), recent thymic emigration (CD31), and the IL‐7 receptor‐α (IL‐7Rα). Serum was analyzed for IL‐7. Alemtuzumab induction produced profound lymphopenia followed by repopulation, during which naive IL‐7Rα⁺CD57^?PD1^? cells progressively became the predominant subset. This did not occur in a comparator group of 10 patients treated with conventional immunosuppression. Serum from depleted patients showed markedly elevated IL‐7 levels posttransplantation. Sorted CD57^?PD1^? cells demonstrated robust proliferation in response to IL‐7, whereas more differentiated cells proliferated poorly. These data suggest that differences in IL‐7‐dependent proliferation is one exploitable mechanism that distinguishes CoB‐sensitive and CoB‐resistant T cell populations to reduce the risk of CoBRR. (ClinicalTrials.gov ‐ NCT00565773.)     

The Effect of Costimulatory and Interleukin 2 Receptor Blockade on Regulatory T Cells in Renal Transplantation

Regulatory T cells (Treg) are critical regulators of immune tolerance. Both IL‐2 and CD28‐CD80/CD86 signaling are critical for CD4⁺CD25⁺FOXP3⁺ Treg survival in mice. Yet, both belatacept (a second‐generation CTLA‐4Ig) and basiliximab (an anti‐CD25 monoclonal antibody) are among the arsenal of current immunotherapies being used in kidney transplant patients. In this study, we explored the direct effect of basiliximab and belatacept on the Tregs in peripheral blood both in the short term and long term and in kidney biopsies of patients with acute rejection. We report that the combined belatacept/basiliximab therapy has no long‐term effect on circulating Tregs when compared to a calcineurin inhibitor (CNI)‐treated group. Moreover, belatacept‐treated patients had a significantly greater number of FOXP3⁺ T cells in graft biopsies during acute rejection as compared to CNI‐treated patients. Finally, it appears that the basiliximab caused a transient loss of both FOXP3⁺ and FOXP3^? CD25⁺ T cells in the circulation in both treatment groups raising important questions about the use of this therapy in tolerance promoting therapeutic protocols.     

Postdepletion Lymphocyte Reconstitution During Belatacept and Rapamycin Treatment in Kidney Transplant Recipients

Belatacept is used to prevent allograft rejection but fails to do so in a sizable minority of patients due to inadequate control of costimulation‐resistant T cells. In this study, we report control of costimulation‐resistant rejection when belatacept was combined with perioperative alemtuzumab‐mediated lymphocyte depletion and rapamycin. To assess the means by which the alemtuzumab, belatacept and rapamycin (ABR) regimen controls belatacept‐resistant rejection, we studied 20 ABR‐treated patients and characterized peripheral lymphocyte phenotype and functional responses to donor, third‐party and viral antigens using flow cytometry, intracellular cytokine staining and carboxyfluorescein succinimidyl ester–based lymphocyte proliferation. Compared with conventional immunosuppression in 10 patients, lymphocyte depletion evoked substantial homeostatic lymphocyte activation balanced by regulatory T and B cell phenotypes. The reconstituted T cell repertoire was enriched for CD28⁺ naïve cells, notably diminished in belatacept‐resistant CD28^? memory subsets and depleted of polyfunctional donor‐specific T cells but able to respond to third‐party and latent herpes viruses. B cell responses were similarly favorable, without alloantibody development and a reduction in memory subsets—changes not seen in conventionally treated patients. The ABR regimen uniquely altered the immune profile, producing a repertoire enriched for CD28⁺ T cells, hyporesponsive to donor alloantigen and competent in its protective immune capabilities. The resulting repertoire was permissive for control of rejection with belatacept monotherapy.

     

T cell exhaustion is associated with antigen abundance and promotes transplant acceptance

Exhaustion of T cells limits their ability to clear chronic infections or eradicate tumors. Here, in the context of transplant, we investigated whether T cell exhaustion occurs and has a role in determining transplant outcome. A peptide/MHC tetramer‐based approach was used to track exhausted CD8⁺ T cells in a male‐to‐female skin transplant model. Transplant of large whole‐tail skins, but not small tail skins (0.8 cm × 0.8 cm), led to exhaustion of anti‐male tetramer⁺ CD8⁺ T cells and subsequently the acceptance of skin grafts. To study CD4⁺ T cell exhaustion, we used the TCR‐transgenic B6 TEa cells that recognize a major transplant antigen I‐Eα from Balb/c mice. TEa cells were adoptively transferred either into B6 recipients that received Balb/c donor skins or into CB6F1 mice that contained an excessive amount of I‐Eα antigen. Adoptively transferred TEa cells in skin‐graft recipients were not exhausted. By contrast, virtually all adoptively transferred TEa cells were exhausted in CB6F1 mice. Those exhausted TEa cells lost ability to reject Balb/c skins upon further transfer into lymphopenic B6.Rag1^?/? mice. Hence, T cell exhaustion develops in the presence of abundant antigen and promotes transplant acceptance. These findings are essential for better understanding the nature of transplant tolerance.     

Regulatory T Cells Are Critical to Tolerance Induction in Presensitized Mouse Transplant Recipients Through Targeting Memory T Cells

Memory T cells are a significant barrier to induction of transplant tolerance. However, reliable means to target alloreactive memory T cells have remained elusive. In this study, presensitization of BALB/c mice with C57BL/6 skin grafts generated a large number of OX40⁺CD44^hieffector/memory T cells and resulted in rapid rejection of donor heart allografts. Recognizing that anti‐OX40L monoclonal antibody (mAb) (α‐OX40L) monotherapy prolonged graft survival through inhibition and apoptosis of memory T cells in presensitized recipients, α‐OX40L was added to the combined treatment protocol of LF15–0195 (LF) and anti‐CD45RB (α‐CD45RB) mAb—a protocol that induced heart allograft tolerance in non‐presensitized recipients but failed to induce tolerance in presensitized recipients. Interestingly, this triple therapy restored donor‐specific heart allograft tolerance in our presensitized model that was associated with induction of tolerogenic dendritic cells and CD4⁺CD25⁺Foxp3⁺ T regulatory cells (Tregs). Of note, CD25⁺ T cell depletion in triple therapy recipients prevented establishment of allograft tolerance. In addition, adoptive transfer of donor‐primed effector/memory T cells into tolerant recipients markedly reduced levels of Tregs and broke tolerance. Our findings indicated that targeting memory T cells, by blocking OX40 costimulation in presensitized recipients was very important to expansion of Tregs, which proved critical to development of tolerance.     

Antigen‐specific CD4+CD25+ T cells induced by locally expressed ICOS‐Ig: the role of Foxp3, Perforin,Granzyme B and IL‐10 ‐ an experimental study

We have previously reported that ICOS‐Ig expressed locally by a PIEC xenograft induces a perigraft cellular accumulation of CD4⁺CD25⁺Foxp3⁺ T cells and specific xenograft prolongation. In the present study we isolated and purified CD4⁺CD25⁺ T cells from ICOS‐Ig secreting PIEC grafts to examine their phenotype and mechanism of xenograft survival using knockout and mutant mice. CD4⁺CD25⁺ T cells isolated from xenografts secreting ICOS‐Ig were analysed by flow cytometry and gene expression by real‐time PCR. Regulatory function was examined by suppression of xenogeneic or allogeneic primed CD4 T cells in vivo. Graft prolongation was shown to be dependent on a pre‐existing Foxp3⁺ Treg, IL‐10, perforin and granzyme B. CD4⁺CD25⁺Foxp3⁺ T cells isolated from xenografts secreting ICOS‐Ig demonstrated a phenotype consistent with nTreg but with a higher expression of CD275 (ICOSL), expression of CD278 (ICOS) and MHC II and loss of CD73. Moreover, these cells were functional and specifically suppressed xenogeinic but not allogeneic primed T cells in vivo.     

Cumulative exposure to CD8+ granzyme Bhi T cells is associated with reduced lung function early after lung transplantation

Outcomes following lung transplant remain suboptimal. This is attributable to variable posttransplant recovery of lung function, and inconsistent degrees of lung function loss after peak function is reached. Granzyme B is elevated in the blood and bronchoalveolar lavage (BAL) in acute rejection. We hypothesized that persistent exposure to T cells high in granzyme B would negatively correlate with lung function. We investigated cumulative exposure measured as the area-under-the-curve (AUC) of CD8⁺ T cell granzyme B^hi cells in the first year posttransplant in both BAL and blood in 24 transplant recipients. We assessed the correlation between cumulative 1-year exposure and FEV₁ slope. There was a negative correlation between 1-year exposure and FEV₁ slope within the first year (r = −0.63; P = .001). This relationship persisted even when adjusted for transplant type, gender, age, rejection, and indication for transplantation. In contrast, no relationship was seen with the 1-year AUC and lung function after 1 year posttransplant. In contrast to the BAL granzyme B^hi levels, granzyme B^hi levels from the blood showed no relationship with lung function. These findings suggest that CD8⁺ T-cell–driven factors are responsible for early improvements in lung function after transplantation.