Analysis of allogeneic and syngeneic bone marrow stromal cell graft survival in the spinal cord

Bone marrow stromal cells (MSC) are attractive candidates for developing cell therapies for central nervous system (CNS) disorders. They can be easily obtained, expanded in culture, and promote modest functional recovery following transplantation into animal models of injured or degenerative CNS. While syngeneic MSC grafts can be used efficiently, achieving long-term survival of allogeneic MSC grafts has been a challenge. We hypothesize that improved graft survival will enhance the functional recovery promoted by MSC. To improve MSC graft survival, we tested two dosages of the immune suppressant cyclosporin A (CsA) in an allogeneic model. Syngeneic transplantation of MSC where cells survive well without immune suppression was used as a control. Sprague-Dawley rats treated with standard dose (n = 12) or high-dose (n = 12) CsA served as allogeneic hosts; Fisher 344 rats (n = 12) served as syngeneic hosts. MSC were derived from transgenic Fisher 344 rats expressing human placental alkaline phosphatase and were grafted into cervical spinal cord. Animals treated with standard dose CsA showed significant decreases in allograft size 4 weeks posttransplantation; high CsA doses yielded significantly better graft survival 4 and 8 weeks posttransplantation compared to standard CsA. As expected, syngeneic MSC transplants showed good graft survival after 4 and 8 weeks. To investigate MSC graft elimination, we analyzed immune cell infiltration and cell death. Macrophage infiltration was high after 1 week in all groups. After 4 weeks, high-dose CsA and syngeneic animals showed significant reductions in macrophages at the graft site. Few T lymphocytes were detected in any group at each time point. Cell death occurred throughout the study; however, little apoptotic activity was detected. Histochemical analysis revealed no evidence of neural differentiation. These results indicate that allogeneic transplantation with appropriate immune suppression permits long-term survival of MSC; thus, both allogeneic and syngeneic strategies could be utilized in devising novel therapies for CNS injury.     

Dexamethasone Prolongs Cardiac Allograft Survival in a Murine Model Through Myeloid-derived Suppressor Cells

Background

Recently, myeloid-derived suppressor cells (MDSCs) have attracted considerable attention because of their cancer-promoting and immunosuppressive effects. The glucocorticoid dexamethasone (Dex) is an important immunosuppressive agent used to treat autoimmune diseases and organ transplant rejection. However, the mechanism by which it modulates the immune system is not completely understood.

Potential T regulatory cell therapy in transplantation: how far have we come and how far can we go?

Graft survival has been lately improved by the introduction of efficient immunosuppressive drugs. However, late graft loss caused by chronic rejection and the side effects of long‐term immunosuppression remain major obstacles for successful transplantation. Operational tolerance, which is defined by the lack of acute and chronic rejection and indefinite graft survival with normal graft function in the absence of continuous immunosuppression, represents an attractive alternative. Nevertheless, tolerance after allogeneic transplantation is commonly considered the ‘mission impossible’ for both immunologists and clinicians. One of the mechanisms involved in tolerance is the suppression of graft‐specific alloreactive T cells, which largely mediate graft rejection, by regulatory T cells (Tregs) or by soluble factors produced by Treg cells. With this review, I will make an effort to collect and describe the significant studies performed in transplanted patients, and not in animal models or in in vitro systems, with the attempt to: (i) understand how tolerance is achieved, (ii) define whether and how Treg cells influence transplant tolerance, (iii) describe the first clinical trials with Treg cells in humans (i.e. how far have we come) and (iv) predict the future of Treg cell‐based therapy in humans (i.e. how far can we go).     

Limited efficacy of rapamycin monotherapy in vascularized composite allotransplantation

BackgroundVascularized composite allotransplantation (VCA) is a novel and life-enhancing procedure to restore a patient's function and/or appearance. Current immunosuppression in VCA recipients is based on calcineurin inhibitor (CNI) therapy that can lead to severe complications, such that inducing immune tolerance is a major goal of VCA research. In contrast to CNI, rapamycin (RPM) is thought to be beneficial to the development of immune tolerance by suppressing T-effector cells (Teffs) and expanding T-regulatory (Treg) cells. However, we found high dose RPM monotherapy prolonged VCA survival by only a few days, leading us to explore the mechanisms responsible.MethodsA mouse orthotopic forelimb transplantation model (BALB/c- > C57BL/6) was established using WT mice, as well as C57BL/6 recipients with conditional deletion of T-bet within their Treg cells. Events in untreated VCA recipients or those receiving RPM or FK506 therapy were analyzed by flow-cytometry, histopathology and real-time qPCR. Results. Therapy with RPM (2 mg/kg/d, p < .005) or FK506 (2 mg/kg/d, p < .005) each prolonged VCA survival. In contrast to FK506, RPM increased the ratio of splenic Treg to Teff cells (p < .05) by suppressing Teff and expanding Treg cells. While the proportion of activated splenic CD4 + Foxp3- T cells expressing IFN-γ were similar in control and RPM-treated groups, RPM decreased the proportions ICOS+ and CD8+ IFN-γ + splenic T cells. However, RPM also downregulated CXCR3+ expression by Tregs, and forelimb allografts had reduced infiltration by CXCR3+ Treg cells. In addition, allograft recipients whose Tregs lacked T-bet underwent accelerated rejection compared to WT mice despite RPM therapy.ConclusionsWe demonstrate that while RPM increased the ratio of Treg to Teff cells and suppressed CD8+ T cell allo-activation, it failed to prevent CD4 Teff cell activation and impaired CXCR3-dependent Treg graft homing, thereby limiting the efficacy of RPM in VCA recipients.     

Allergic Conjunctivitis Renders CD4+ T Cells Resistant to T Regulatory Cells and Exacerbates Corneal Allograft Rejection

Allergic diseases rob corneal allografts of immune privilege and increase immune rejection. Corneal allograft rejection in BALB/c allergic hosts was analyzed using a short ragweed (SWR) pollen model of allergic conjunctivitis. Allergic conjunctivitis did not induce exaggerated T‐cell responses to donor C57BL/6 (B6) alloantigens or stimulate cytotoxic T lymphocyte (CTL) responses. Allergic conjunctivitis did affect T regulatory cells (Tregs) that support graft survival. Exogenous IL‐4, but not IL‐5 or IL‐13, prevented Treg suppression of CD4⁺ effector T cells isolated from naïve mice. However, mice with allergic conjunctivitis developed Tregs that suppressed CD4⁺ effector T‐cell proliferation. In addition, IL‐4 did not inhibit Treg suppression of IL‐4Rα^?/? CD4⁺ T‐cell responses, suggesting that IL‐4 rendered effector T cells resistant to Tregs. SRW‐sensitized IL‐4Rα^?/? mice displayed the same 50% graft survival as nonallergic WT mice, that was significantly less than the 100% rejection that occurred in allergic WT hosts, supporting the role of IL‐4 in the abrogation of immune privilege. Moreover, exacerbation of corneal allograft rejection in allergic mice was reversed by administering anti‐IL‐4 antibody. Thus, allergy‐induced exacerbation of corneal graft rejection is due to the production of IL‐4, which renders effector T cells resistant to Treg suppression of alloimmune responses.     

Graft produced interleukin-6 functions as a danger signal and promotes rejection after transplantation

BACKGROUND: Interleukin (IL)-6 is a pleiotropic cytokine that functions in both the innate and adaptive immune responses. However, the role of IL-6 in allograft rejection remains poorly understood. METHODS: In this study, we demonstrate a critical role for graft-produced IL-6 in allograft rejection in a murine model of cardiac allograft transplantation. RESULTS: The results show that IL-6-deficient grafts transplanted into allogeneic wild-type recipients have significantly prolonged survival, approximately three times the survival time of wild-type controls. In contrast, allogeneic cardiac transplants into IL-6-deficient recipients do not have prolonged graft survival, indicating that donor graft cells are the relevant source of IL-6. Our investigation of potential mechanisms shows that graft-produced IL-6 promotes the activation of peripheral CD4 and CD8 T cells. Furthermore, we show that IL-6 deficiency prolongs graft survival only in the presence of CD25+ T cells that have a phenotype consistent with regulatory T cells. Interestingly, IL-6 production by the graft is triggered by antigen-independent innate immune mechanisms. CONCLUSIONS: Thus, our results suggest the paradigm that graft rejection versus tolerance is determined by a balance between the activation of effector T cells versus immune suppression by regulatory T cells, and that after transplantation, IL-6 functions as a systemic danger signal that overcomes constitutive immune suppression mediated by regulatory T cells and promotes the activation of effector T cells.     

The relationship between T-cell infiltration in biopsy proven acute T-cell mediated rejection with allograft function and response to therapy: A retrospective study

AimTo evaluate the degree of CD3, CD20, Th17, and Tregs infiltration in kidney biopsy of the patients with acute cellular rejection and the possible relation with graft outcome.Materials and methodsIn this retrospective study, fifty patients with Acute T Cell-Mediated Rejection (ATCMR) were enrolled. Previous and one year clinical follow-up data were collected. The kidney specimens were evaluated for infiltration of CD3, CD20, FOXP3, and Th17 with IHC. According to the serum creatinine level in one-year follow-up of the patients after rejection therapy and function of the transplanted organ from the day admitted into the hospital, they were respectively categorized in Stable graft function versus impaired graft function; appropriate response to treatment versus failure to response.ResultsTreg (P = 0.96) and Th17 (P = 0.24) cells were more in the unstable group than the stable group, but the difference wasn't significant. On the other hand, the FOXP3/Th17 ratio was higher in the stable group (P = 0.22). Moreover Treg (P = 0.1) and Th17 (P = 0.15) were higher in failure to response group, but FOXP3/Th17 was higher in proper response group (P = 0.8).ConclusionFrom the results, it can be concluded that TH17 infiltration has a more significant effect on graft outcome and response to rejection therapy.     

BEZ235 Prolongs Murine Cardiac Allograft Survival Through the Autophagy Pathway

BackgroundBEZ235, a dual PI3K/mTOR inhibitor, has shown a critical impact in the treatment of cancers, with the ability to induce autophagy. However, the effects of BEZ235 in heart transplant have been rarely investigated. The aim of this study was to evaluate the potency of BEZ235 in cardiac allograft survival.MethodsBEZ235 was administered during the perioperative period of syngeneic or allogeneic heart transplant to assess survival time. Next, the autophagy signaling pathway and the proinflammatory cytokines were analyzed. Furthermore, a cardiomyocytes-specific ATG5 gene-ablated mouse was used to confirm the results.ResultsBEZ235 treatment significantly prolonged the survival of the cardiac graft and reduced the infiltration of inflammatory cells. The expression levels of autophagy proteins were increased in the BEZ235 treatment group compared to the control group, but the therapeutic effect of BEZ235 was weakened in the cardiomyocytes-specific ATG5 gene-ablated mice. Moreover, BEZ235 significantly downregulated the expression of IL-1β, IL-2, and TNF-α.ConclusionsIt seems BEZ235 could induce autophagy and prolonged murine cardiac allograft survival in a mechanism that involved the autophagy pathway and changed multiple inflammatory factors. This study has proposed a theoretical foundation for the strong connection between mTOR-induced autophagy and heart transplant.     

Prevention of Allograft Rejection by Use of Regulatory T Cells With an MHC‐Specific Chimeric Antigen Receptor

CD4⁺CD25^highFOXP3⁺ regulatory T cells (Tregs) are involved in graft‐specific tolerance after solid organ transplantation. However, adoptive transfer of polyspecific Tregs alone is insufficient to prevent graft rejection even in rodent models, indicating that graft‐specific Tregs are required. We developed a highly specific chimeric antigen receptor that recognizes the HLA molecule A*02 (referred to as A2‐CAR). Transduction into natural regulatory T cells (nTregs) changes the specificity of the nTregs without alteration of their regulatory phenotype and epigenetic stability. Activation of nTregs via the A2‐CAR induced proliferation and enhanced the suppressor function of modified nTregs. Compared with nTregs, A2‐CAR Tregs exhibited superior control of strong allospecific immune responses in vitro and in humanized mouse models. A2‐CAR Tregs completely prevented rejection of allogeneic target cells and tissues in immune reconstituted humanized mice in the absence of any immunosuppression. Therefore, these modified cells have great potential for incorporation into clinical trials of Treg‐supported weaning after allogeneic transplantation.     

Ex-vivo expanded baboon CD4+ CD25 Hi Treg cells suppress baboon anti-pig T and B cell immune response

Singh AK, Seavey CN, Horvath KA, Mohiuddin MM. Ex‐vivo expanded baboon CD4⁺ CD25^Hi Treg cells suppress baboon anti‐pig T and B cell immune response. Xenotransplantation 2012; 19: 102–111. © 2012 John Wiley & Sons A/S. Abstract: Background: CD4⁺ CD25⁺ FoxP3⁺ regulatory T (Treg) cells play an important role in regulating immune responses. A very small number of Treg cells are present in peripheral blood and lymphoid organs, but due to their ability to suppress the immune response, they have a high potential for immunotherapy in clinics. Successful ex‐vivo expansion of naturally occurring CD4⁺ CD25⁺ T cells has been achieved after TCR stimulation in the presence of T cell growth factors. In this study, we evaluated the role of these Treg cells in suppressing proliferative response of baboon T and B cells to pig xenoantigens. Methods: Naturally occurring baboon CD4⁺ CD25⁺ regulatory T cells (nTreg) were sorted from peripheral blood and expanded in the presence of either anti‐CD3/CD28 beads or irradiated pig peripheral blood mononuclear cells with IL‐2. Treg cells were also enriched directly from CD4⁺ T cells cultured in the presence of rapamycin (0.1–10 nm ). Mixed lymphocyte culture and polyclonal B cell stimulation with ex‐vivo Treg cells were performed to assess the function of ex‐vivo expanded Treg cells. Results: The nTreg cells were expanded to more than 200‐fold in 4 weeks and retained all the nTreg cell phenotypic characteristics, including high levels of FoxP3 expression. 2‐fold increase in enrichment of CD4⁺ CD25⁺ FoxP3⁺ Treg cells from CD4⁺ cells was observed with rapamycin compared to cultures without rapamycin. The ex‐vivo expanded Treg cells obtained from both methods were able to suppress the baboon anti‐porcine xenogeneic T and B cell immune response in‐vitro efficiently (more than 90% suppression at 1 : 1 ratio of T regulatory cells: T effector cells), and their suppression potential was retained even at 1 : 256 ratio. However, freshly isolated nTreg cells had only 70% suppression at 1 : 1 ratio, and their suppressive ability was reduced to ≤50% at 1 : 16 ratio. Furthermore, we have found that ex‐vivo expanded Treg can also suppress the proliferation of B cells after polyclonal stimulation. Forty to 50 percent reduction in B cell proliferation was observed when ex‐vivo expanded Treg cells were added to the culture at a 1 : 1 ratio. The addition of CD4⁺ CD25^Neg cells however induced vigorous proliferation. Conclusion: Ex‐vivo expanded CD4⁺ CD25⁺ FoxP3⁺ Treg cells can be used to efficiently suppress xenogeneic immune responses by inhibiting T and B cell proliferation. These ex‐vivo expanded Treg cells may also be used with other immunosuppressive agents to overcome xenograft rejection in preclinical xenotransplantation models.     

Absolute of CD4(+)CD25(+)FOXP3(+) regulatory T-cell count rather than its ratio in peripheral blood is related to long-term survival of renal allografts

CD4⁺CD25⁺Foxp3⁺ regulatory T cells (Tregs) may reflect the immune status of kidney transplant (KTx) recipients. Since individual KTx recipients show different lymphocyte counts, we hypothesized that the Treg absolute count rather than its peripheral ratio was more related to long-term survival. We enrolled 42 patients with more than 5-year KTx survival: 32 patients with stable graft function and 10 suffering chronic rejection (CR group). The stable group was divided into four subgroups according to graft survival time: subgroups A (5-6 years); B (6-7 years); C (7-8 years); and D (>8 years). Healthy volunteers were enrolled as controls. We compared the peripheral ratio and absolute count of CD4⁺, CD4⁺CD25⁺, and CD4⁺CD25⁺Foxp3⁺ Tregs. Treg peripheral ratio was not significantly different among the three groups. However, the Treg absolute count was higher among the stable than the CR group (P < .01). Meanwhile, both Treg ratios and absolute counts were altered with renal graft survival. Treg absolute count in subgroups B and C were not only higher than that in subgroup A (P < .05), but also was significantly higher than that in the CR control group or (P < .05). Interestingly, both the Treg ratio and absolute count in subgroup D were lower than those in subgroups B and C (P < .05). Treg quantitation, which alters with graft survival time, may contribute to long-term acceptance of renal allografts. Compared to Treg peripheral ratio, the absolute count may be a superior index to estimate KTx recipient immune status.     

Hepatic Stellate Cells Induce Immunotolerance of Islet Allografts

Activated hepatic stellate cells (HSCs) possess strong immune inhibitory activity. The present study highlighted the protective role of HSCs in islet transplantation. Recipients were randomly divided into 4 groups: a diabetic group, an HSC-alone group, an islet-alone transplant group, and a cotransplant group. Graft survival was compared among the 4 groups. Serum transforming growth factor β (TGFβ), tumor necrosis factor α, interleukin-1β, and interferon gamma expression levels were measured. The infiltration of lymphocytes was observed via hematoxylin and eosin staining, and immunohistochemical examinations were performed. Results showed that allogeneic HSCs protect islet allografts better than syngeneic HSCs. There was significant prolonged graft survival and a higher level of TGFβ in the cotransplant group (P < .01). The infiltration of lymphocytes in the cotransplant group was notably less than in the islet-alone group (P < .01). The formation of desmin-positive HSC packages was detected in the cotransplant group. In conclusion, allogeneic HSCs can better prolong the survival of islet allografts by stimulating TGFβ expression and forming a biological capsule around the graft.     

Adoptive transfer with in vitro expanded human regulatory T cells protects against porcine islet xenograft rejection via interleukin-10 in humanized mice

T cell-mediated rejection remains a barrier to the clinical application of islet xenotransplantation. Regulatory T cells (Treg) regulate immune responses by suppressing effector T cells. This study aimed to determine the ability of human Treg to prevent islet xenograft rejection and the mechanism(s) involved. Neonatal porcine islet transplanted NOD-SCID IL2rγ(-/-) mice received human peripheral blood mononuclear cells (PBMC) with in vitro expanded autologous Treg in the absence or presence of anti-human interleukin-10 (IL-10) monoclonal antibody. In addition, human PBMC-reconstituted recipient mice received recombinant human IL-10 (rhIL-10). Adoptive transfer with expanded autologous Treg prevented islet xenograft rejection in human PBMC-reconstituted mice by inhibiting graft infiltration of effector cells and their function. Neutralization of human IL-10 shortened xenograft survival in mice receiving human PBMC and Treg. In addition, rhIL-10 treatment led to prolonged xenograft survival in human PBMC-reconstituted mice. This study demonstrates the ability of human Treg to prevent T-cell effector function and the importance of IL-10 in this response. In vitro Treg expansion was a simple and effective strategy for generating autologous Treg and highlighted a potential adoptive Treg cell therapy to suppress antigraft T-cell responses and reduce the requirement for immunosuppression in islet xenotransplantation.     

Role of cold ischemia in acute rejection: characterization of a humoral-like acute rejection in experimental renal transplantation

The aim of the study was to characterize the role of cold ischemia in the process of acute rejection using an experimental renal transplant model. Syngeneic renal transplants were performed between Wistar Agouti rats and allogeneic grafts using Wistar-Agouti rats as recipients of Brown-Norway kidneys. For cold ischemia (CI), kidneys were preserved in Euro-Collins (4 degrees C/ 2.5 hours). Rats were bilaterally nephrectomized at the moment of renal transplant and did not receive any immunosuppressant. The groups were NoAR (n = 6): immediate syngeneic transplant; CI-NoAR (n = 6): syngeneic transplant with CI; AR (n = 13): immediate allogeneic graft; CI-AR (n = 6): allogeneic graft with CI. Allogeneic rats were followed for the survival study. Syngeneic rats, with mean survival time beyond 6 months, were sacrificed on the day 7 to compare grafts with those in the allogeneic groups. H&E- and PAS-stained grafts were evaluated using the Banff criteria. Tissue INF-gamma and TNF-alpha were quantified by RT-real time-PCR on the kidney grafts. Renal insufficiency did not appear in the NoAR group, but it did from the posttransplant day 5 in both acute rejection groups. While NoAR kidneys showed well-conserved renal architecture, then AR group displayed variable degrees of tubular necrosis with scarce cellular infiltration, interstitial hemorrhage, vascular damage with fibrinoid necrosis, perivascular edema, and nuclear disruption. Cold ischemia in rejecting animals increased the mortality rate due to renal insufficiency and accelerated acute rejection. Independently of CI, the proinflammatory cytokines TNF-alpha and INF-gamma were increased in both rejection groups. In conclusion, addition of CI overactivates the acute rejection process via a humoral component.     

Phenotyping of ex vivo propagated graft-infiltrating cells-a tool to monitor rejection in the early post-operative period

Objective and fast methods to diagnose rejection after organ transplantation are needed. In the present study, the ex vivo propagation technique was evaluated for its ability to detect rejection at two different time-points after experimental heart transplantation. Syngeneic and allogeneic heterotopic heart transplantations were performed using inbred rat strains. After 6 or 15 days, cardiac graft biopsies were put in culture and infiltrating cells isolated by the ex vivo propagation technique. The isolated cells were counted and phenotyped by flow cytometry. In parallel, graft sections were analysed with regard to morphology and the presence of infiltrating cells as determined by immunohistochemical stainings. On day 15 after transplantation, the number of cells possible to isolate through ex vivo propagation reflected the morphological changes of the graft, i.e. considerably more cells were obtained from allogeneic transplants undergoing rejection (1052 +/- 205) than from allogeneic grafts under cyclosporine protection (513 +/- 135; p < 0.05) or from syngeneic grafts (378 +/- 87; p < 0.01). Six days after transplantation the allogeneic grafts were strongly rejected with massive cellular infiltration, still there was no difference between allogeneic and syngeneic grafts as to the number of ex vivo propagated cells. However, the proportion of IL-2-receptor expressing T lymphocytes was increased (15.4 +/- 1.8% vs. 9.5 +/- 1.4%; p < 0.05) and the CD4/CD8 ratio reduced (1.0 +/- 0.1 vs. 2.8 +/- 0.2; p < 0.001) in the allogeneic group as compared with the syngeneic. We conclude that the ex vivo propagation technique can be used to distinguish rejection from non-rejection both early and later after transplantation, provided that not just cell counting but also phenotyping of the graft-infiltrating cells is performed.     

In vivo induction of myeloid suppressor cells and CD4(+)Foxp3(+) T regulatory cells prolongs skin allograft survival in mice

Natural CD4(+)Foxp3(+) T regulatory (Treg) cells can promote transplantation acceptance across major histocompatibility complex (MHC) barriers, while myeloid-derived suppressor cells (MDSCs) inhibit effector T-cell responses in tumor-bearing mice. One outstanding issue is whether combining the potent suppressive function of MDSCs with that of Treg cells might synergistically favor graft tolerance. In the present study, we evaluated the therapeutic potential of MDSCs and natural Treg cells in promoting allograft tolerance in mice by utilizing immunomodulatory agents to expand these cells in vivo. Upon administration of recombinant human granulocyte-colony stimulating factor (G-CSF; Neupogen), or interleukin-2 complex (IL-2C), Gr-1(+)CD11b(+) MDSCs or CD4(+)Foxp3(+) Treg cells were respectively induced at a high frequency in the peripheral lymphoid compartments of treated mice. Interestingly, induced MDSCs exhibited a more potent suppressive function in vitro when compared to MDSCs from naive mice. Furthermore, in vivo coadministration of Neupogen and IL-2C induced MDSCs at percentages that were higher than those seen when either agent was administered alone, suggesting an additive effect of the two drugs. Although treatment with either IL-2C or Neupogen led to a significant delay of MHC class II disparate allogeneic donor skin rejection, the combinatorial treatment was superior to either alone. Importantly, histological assessment of surviving grafts revealed intact morphology and minimal infiltrates at 60 days posttransplant. Collectively, our findings demonstrate that concurrent induction of MDSCs and Tregs is efficacious in downmodulating alloreactive T-cell responses in a synergistic manner and highlight the therapeutic potential of these naturally occurring suppressive leukocytes to promote transplantation tolerance.     

Adoptive transfer of double negative T regulatory cells induces B-cell death in vivo and alters rejection pattern of rat-to-mouse heart transplantation

Abstract: Background: Antibody‐mediated hyperacute and acute graft rejection are major obstacles in achieving long‐term graft survival in xenotransplantation. It is well documented that regulatory T (Treg) cells play a very important role in regulating immune responses to self and non‐self antigens. Our previous studies have shown that TCRαβ⁺CD3⁺CD4^?CD8^? (double negative, DN)‐Treg cells can suppress anti‐donor T‐cell responses and prolong graft survival in allo‐ and xenotransplantation models. We have demonstrated that DN‐Treg cells can induce B‐cell apoptosis in vitro through a perforin‐dependent pathway. Methods: B6 mice received rat heart grafts, followed by 14 days of LF15‐0195 treatment. Some mice received Lewis rat cell activated DN‐Treg cells after LF treatment. DN‐Treg cells, purified from perforin^?/? mice and from B6 mice pre‐immunized with third party rat cells, were used as controls. Results: In this study, we investigated the possibility that adoptive transfer of xenoreactive DN‐Treg cells could suppress B cells in vivo, thus prolonging xenograft survival. We found that apoptotic death of B cells significantly increased after adoptive transfer of DN‐Treg cells. In addition, anti‐donor IgG subtypes were significantly inhibited in the DN‐Treg cell‐treated group, in which the rejection pattern was altered towards cellular‐mediated rejection rather than antibody‐mediated acute vascular rejection. However, perforin‐deficient DN‐Treg cells failed to induce B‐cell death and to prolong heart graft survival, indicating a perforin‐dependent mechanism contributes to B‐cell death in vivo. Conclusions: This study suggests that adoptive transfer of xenoreactive DN‐Treg cells can inhibit B‐cell responses in vivo. DN‐Treg cells may be valuable in controlling B‐cell responses in xenotransplantation.     

Leptin Modulates Allograft Survival by Favoring a Th2 and a Regulatory Immune Profile

Leptin, an adipose‐secreted hormone, links metabolism and immunity. Our aim was to determine whether leptin affects the alloimmune response. We used an allogeneic skin transplant model as a means to analyze the allograft immune response in Lep^ob/ob and wild‐type mice. Leptin deficiency results in an increased frequency of Treg and Th2 cells and a prolonged graft survival. These effects of leptin deficiency indicate the importance of leptin and obesity in modulating the allograft immune responses. Our data suggest a possible explanation for the increased susceptibility of hyperleptinemic obese patients to acute and chronic graft rejection.     

Therapeutic efficacy of extracellular vesicles to suppress allograft rejection in preclinical kidney transplantation models: A systematic review and meta-analysis

BackgroundKidney transplantation is the optimal treatment of end-stage renal disease. Extracellular vesicles (EVs) have tremendous therapeutic potential, but their role in modulating immune responses in kidney transplantation remains unclear.MethodsWe performed a systematic review and meta-analysis to investigate the therapeutic efficacy of EVs in preclinical kidney transplant models. Outcomes for meta-analysis were graft survival and renal function. Subgroup analysis was conducted between immune cell derived EVs (immune cell-EVs) and mesenchymal stromal cell derived EVs (MSC-EVs).ResultsSeven studies published from 2013 to 2021 were included. The overall effects showed that EVs had a positive role in prolonging allograft survival (standardized mean difference (SMD) = 2.00; 95% confidence interval (CI), 0.79 to 3.21; P < 0.01; I² = 94%), reducing serum creatinine (SCr) (SMD = -2.19; 95%CI, ?3.35 to ?1.04; P < 0.01; I² = 93%) and blood urea nitrogen (BUN) concentrations (SMD = -1.69; 95%CI, ?2.98 to ?0.40; P = 0.01; I² = 94%). Subgroup analyses indicated that only immune cell-EVs significantly prolonged graft survival and improve renal function but not MSC-EVs.ConclusionsEVs are promising candidates to suppress allograft rejection and improve kidney transplant outcome. Immune cell-EVs showed their superiority over MSC-EVs in prolonging graft survival and improving renal function. For interpretation of the outcomes, additional studies are needed to validate these findings.