Liver Transplant Tolerance Research Articles

Human alloreactive-regulatory T cell profile after ex vivo expansion using CD40L-stimulated B cells or monocyte-derived dendritic cells

Abstract Alloreactive regulatory T cells (arTregs) expanded with CD40L-stimulated B cells (sBcs) are being tested clinically to induce tolerance and control rejection in liver and kidney transplantation. In research settings, allogeneic monocyte-derived DCs (moDCs) have been shown to expand arTregs. We directly compared the efficacy of sBcs and cytokine-matured moDCs in expanding arTregs, and the phenotype of sBc-stimulated arTregs (sBc-arTregs) and moDC-stimulated arTregs (moDC-arTregs). arTregs expanded twice as much after stimulation with moDCs than with sBcs. Neither sBcs nor moDCs produced much TNFα, IL-1α, IL-1β, IL-6, IL-12P70, or IL-23 that can destabilize Tregs. sBc- and moDC-arTregs expressed similar percentages of Treg-defining markers - FOXP3, HELIOS, CD25, CD27, and CD62L. moDC-arTregs had slightly elevated FOXP3 and CD62L MFI. Compared to primary Tregs, sBc- and moDC-arTregs expressed similar or decreased levels of TBX21, GATA3, and RORC mRNA. However, greater percentages of moDC-arTregs produced IFNγ, IL-4, and IL-17A, and a greater percentage of sBc-arTregs produced IL-17A, but far less than those percentages produced by similarly stimulated conventional CD4+ T cells (Tconvs). The majority of cytokine-expressing arTregs were FOXP3+. Percentages of CXCR3 and CCR4, but not CCR6, increased after sBc and moDC stimulation. Using a 770-gene Nanostring panel and unsupervised clustering, sBc- and moDCs-arTregs were found to be mostly similar, but distinct from primary Tregs and further separated from Tconvs. Thus, arTregs expanded by sBcs or moDCs are phenotypically similar, likely stable, and had features that may enable them to traffic to inflammatory sites, e.g. transplanted organs undergoing alloimmune attack.

PDL1Ig gene-loaded BMSCs Induce liver transplantation immune tolerance.

We investigated the effects of BMSCs, which was modified by programmed death ligand-1 immunoglobulin (PDL1Ig) gene, on the immunological rejection of orthotopic liver transplantation (OLT) in rats. Rat BMSCs were cultured and modified by recombinant adenovirus pAdEasy-1/PDL1Ig for 72h. The total protein was extracted, and the protein expression of PDL1Ig after transfection was detected by Western blotting. Mixed lymphocyte reaction was applied to detect the inhibitory effect of BMSCs, including pre-transfection and post-transfection, on the cell activity of T lymphocytes in peripheral blood. The male Wistar rats were used as donors, and the male Sprague-Dawley (SD) rats were used as recipients. The improved cuff method was computed in OLT to establish the rat orthotopic liver transplantation model of acute rejection. The rats were randomly divided into 4 groups, including a control group (10 pairs), BMSCs treatment group (10 pairs), BMSCs/GFP (green fluorescent protein) treatment group (10 pairs), and BMSCs/PDL1Ig treatment group (10 pairs). 5 rats in each group were randomly collected and euthanized at day 7 after the operation. The peripheral blood was gathered to detect levels of 3 types of cytokines, including interferon gamma (IFN-g), interleukin-4 (IL-4), and IL-2. In addition, the liver function of rats was checked, and the pathological changes of liver transplantation were observed under the optical microscope. The remaining five rats in every group were used for measuring the survival situation and survival time. After BMSCs were modified by a recombinant adenovirus pAdEasy-1/PDL1Ig for 72h, the expression of PDL1Ig in the BMSCs was detected. The inhibitory effects of BMSCs/PDL1Ig on the cell activity of lymphocytes were stronger than that of BMSCs/GFP. The levels of IL-4 in BMSCs/PDL1Ig group were significantly higher than those in the other 3 groups, and the levels of IFN-g and IL-2 were significantly decreased (p<0.05). The liver function of 4 groups was measured at day 7 after transplantation. Our results showed that the liver function in BMSCs/PDL1Ig group was most significantly improved, and the levels of ALT, AST, and TBil almost recovered to normal. The differences were statistically significant compared to the control group, BMSCs treatment group, and BMSCs/GFP treatment group. Results of pathological examination of liver tissue showed that the control group underwent severe rejection of liver transplantation. The BMSCs treatment group and BMSCs/GFP treatment group also rejected the liver transplantation, but the degree was lighter when compared to the control group. In addition, there was almost no rejection of liver transplantation in the BMSCs/PDL1Ig group. The recipient survival time of most rats was more than 100d, which was significantly longer than the other 3 groups (p<0.05). PDL1Ig-modified BMSCs can inhibit the rejection of liver transplantation in rats, induce the formation of the immune tolerance of liver transplantation, and the effect was more significant than that of BMSCs alone.

Potential Beneficial Effects of Cytomegalovirus Infection after Transplantation.

Cytomegalovirus (CMV) infection can cause significant complications after transplantation, but recent emerging data suggest that CMV may paradoxically also exert beneficial effects in two specific allogeneic transplant settings. These potential benefits have been underappreciated and are therefore highlighted in this review. First, after allogeneic hematopoietic stem cell transplantation (HSCT) for acute myeloid leukemia (AML) using T-cell and natural killer (NK) cell-replete grafts, CMV reactivation is associated with protection from leukemic relapse. This association was not observed for other hematologic malignancies. This anti-leukemic effect might be mediated by CMV-driven expansion of donor-derived memory-like NKG2C+ NK and Vδ2negγδ T-cells. Donor-derived NK cells probably recognize recipient leukemic blasts by engagement of NKG2C with HLA-E and/or by the lack of donor (self) HLA molecules. Vδ2negγδ T cells probably recognize as yet unidentified antigens on leukemic blasts via their TCR. Second, immunological imprints of CMV infection, such as expanded numbers of Vδ2negγδ T cells and terminally differentiated TCRαβ+ T cells, as well as enhanced NKG2C gene expression in peripheral blood of operationally tolerant liver transplant patients, suggest that CMV infection or reactivation may be associated with liver graft acceptance. Mechanistically, poor alloreactivity of CMV-induced terminally differentiated TCRαβ+ T cells and CMV-induced IFN-driven adaptive immune resistance mechanisms in liver grafts may be involved. In conclusion, direct associations indicate that CMV reactivation may protect against AML relapse after allogeneic HSCT, and indirect associations suggest that CMV infection may promote allograft acceptance after liver transplantation. The causative mechanisms need further investigations, but are probably related to the profound and sustained imprint of CMV infection on the immune system.

Graft-infiltrating PD-L1hi cross-dressed dendritic cells regulate antidonor T cell responses in mouse liver transplant tolerance.

These findings suggest that graft-infiltrating PD-L1hi CD-DCs may play a key role in the regulation of alloimmunity and in the induction of liver transplant tolerance. (Hepatology 2018;67:1499-1515).

Regulatory T-cell therapy in liver transplantation.

Modern immunosuppression drug regimens have produced excellent short-term survival after liver transplantation but it is generally accepted that the side effects of these medications remain a significant contributing factor for less satisfactory long term outcomes. The liver has unique tolerogenic properties as evidenced by the higher rates of operational tolerance seen in liver transplant recipients compared to other solid organ transplants, and therefore, liver transplantation offers an attractive setting in which to study tolerizing therapies. CD4+ CD25+ FOXP3+ regulatory T cells (Tregs) are crucial for maintenance of self-tolerance and prevention of autoimmune disease and are therefore an appealing potential candidate for use as a tolerizing cell therapy. In this review, we summarize the evidence from drug withdrawal trials of spontaneous operational tolerance in liver transplantation, the unique immunology of the hepatic microenvironment, the evidence for the use of CD4+ CD25+ FOXP3+ regulatory T cells as a tolerance inducing therapy in liver transplantation and the challenges in producing clinical grade Treg cell products.

Differential profile of activated regulatory T cell subsets and microRNAs in tolerant liver transplant recipients.

Regulatory T cells (Tregs) play a potential role in operational tolerance in liver transplantation (LT) patients, and microRNAs (miRNAs) are known to be involved in immunological responses and tolerance. Thus, we analyzed the implication of different peripheral blood Treg subsets and miRNAs on LT tolerance in 24 tolerant (Tol) and 23 non-tolerant (non-Tol) LT recipients by cellular, genetic, and epigenetic approximation. Non-Tol patients had a lower demethylation rate of the forkhead box P3 (FOXP3) regulatory T cell-specific demethylated region (TSDR) than Tol patients that correlated with the frequency of circulating Tregs. Tol patients presented a different signature of Treg subset markers compared with non-Tol patients with increased expression of HELIOS and FOXP3 and a higher proportion of latency-associated peptide (LAP)+ Tregs and CD45RA- human leukocyte antigen D related (HLA-DR)+ activated effector-memory Tregs. The expression of miR95, miR24, miR31, miR146a, and miR155 was higher in Tol than in non-Tol patients and was positively correlated with activated Treg markers. In conclusion, these data suggest that activated effector-memory Tregs and a TSDR-demethylation state of Tregs may play a role in the complex system of regulation of LT tolerance. In addition, we describe a set of miRNAs differentially expressed in human LT Tol patients providing suggestive evidence that miRNAs are implied in the preservation of self-tolerance as mediated by Tregs. Liver Transplantation 23 933-945 2017 AASLD.

Graft-Infiltrating PD-L1hi Cross-Dressed Dendritic Cells Subvert Donor-Reactive T Cell Responses in Mouse Liver Transplant Tolerance

Introduction: The mechanisms underlying “spontaneous” liver allograft tolerance remain unclear. Given recent reports of the key role of host dendritic cells (DCs) in mouse spleens that express intact donor MHC (cross-dressing) in the instigation of graft rejection[1][2] we investigated the role of cross-dressed (CD) DCs in mouse liver transplantation tolerance. Methods: Liver allografts from C57BL/6 (B6; H-2b) or B6 SJL CD45.1 mice to C3H/HeJ (C3H; H-2 k) recipients were performed. In this combination, unlike heart, skin or kidney allografts, > 90% of liver grafts are accepted without immunosuppressive therapy[3]. Graft non-parenchymal cells (NPC) and splenocytes were examined by flow cytometry and imaging cytometry on post-operative days (POD) 1, 3, 7, 14, 30, and 300. Mixed leukocyte reactions (CFSE-MLR) were also performed to assess DC function. Results: Infiltration by recipient DCs (lineage (−), CD11c (+)) in liver allografts peaked on POD 7, while donor DCs in liver grafts gradually disappeared and could not be detected by POD 7. Interestingly, more than half of the graft-infiltrating recipient DCs at that time displayed donor MHC-I, indicating cross-dressing; these DCs persisted in the graft (approx. 20 % of recipient DC) at least until POD 300. In contrast, only a very minor fraction of cross-dressed DCs (CD-DCs) (0-2%) were detected in the spleen at any time point. Control staining for donor MHC-I using naïve C3H mouse liver NPC or splenocytes was negative. Moreover, especially on POD 7, and persisting until POD 300, CD-DCs isolated from liver grafts expressed higher levels of T cell inhibitory programed death ligand 1 (PD-L1) compared to non CD-DCs (nCD-DCs). Importantly, unlike nCD-DCs, CD-DCs from liver grafts did not stimulate proliferation of allo-reactive donor T cells and markedly suppressed donor-reactive host T cell proliferation in CFSE-MLR. Conclusions: A large proportion of CD recipient DCs with capacity to subvert donor-reactive host T cell responses are evident in liver allografts early post-transplant. Moreover, cross-dressing by graft-infiltrating DCs that also express high levels of PD-L1, persists indefinitely. This suggests that graft-infiltrating host CD-DC may play a key role in regulation of alloimmunity and the promotion of donor-specific liver transplant tolerance.Figure 1: Marked reduction in donor dendritic cells (DCs) and emergence of cross-dressed host DCs in the graft after liver allo-transplantaion. (A) (B) Graft and spleen DCs were analyzed using donor recipient MHC class 1 (A) or CD45.1/ CD 45.2 antibodies (B) after B6 WT graft to C3H recipient transplatation. Data shown are representative data from 3-5 mice.Figure 2: Infiltrating host DCs and CD-DCs peak on 7 POD and PD-L1high CD-DCs persist in the graft for at least 300 POD. (A) Chronological changes in (left) the incidence of total DC and (right) the incidence of CD-DC (as % total DC) in the graft. (B) Expression of PD-L1 by CD-DCs and non (n)CD-DCs in the graft. PD-L1 expression by CD-DCs was significally higher than of nCD-DC, especially on 7POD. 3-6 mice per time point. *: P<0.05, ***: P<0.001Figure 3: Liver CD-DCs markedly sppress donor-reactive host T cell proliferation in CFSE-MLR Spleen (Sp) or liver CD-DCs or nCD-DCs isolated from graft recipients on 7 POD were co- cultured with responder (C3H (host) spleen T cells) and stimulators (B6 (donor) spleen DC) in CFSE-MLR (T cells: DC (each) =10:1). Liver CD-DCs, markedly suppressed host T cell proliferation. % T cell proliferation in indicated. Data are representative of 3 experiments that gave similar results.

Mass cytometry reveals a distinct immunoprofile of operational tolerance in pediatric liver transplantation

Long-term IS in transplant patients has significant morbidity, poorer quality of life, and substantial economic costs. TOL, defined as graft acceptance without functional impairment in the absence of IS, has been achieved in some pediatric LT recipients. Using mass cytometry, peripheral blood immunotyping was performed to characterize differences between tolerant patients and patients who are stable on single-agent IS. Single-cell mass cytometry was performed using blood samples from a single-center pediatric LT population of operationally tolerant patients to comprehensively characterize the immune cell populations in the tolerant state compared with patients on chronic low-dose IS. Specific T-cell populations of interest were confirmed by flow cytometry. This high-dimensional phenotypic analysis revealed distinct immunoprofiles between transplant populations as well as a CD4+ TOT (CD4+ CD5+ CD25+ CD38-/lo CD45RA) that correlates with tolerance in pediatric LT recipients. In TOL patients, the TOT was significantly increased as compared to patients stable on low levels of IS. This TOT cell was confirmed by flow cytometry and is distinct from classic Treg cells. These results demonstrate the power of mass cytometry to discover significant immune cell signatures that have diagnostic potential.

Mechanisms and Strategies for Tolerance in Liver Transplantation

Although there has been improvement in immunosuppressive therapy in preventing acute rejection following liver transplantation, there is less progress in reducing morbidity and mortality related to immunosuppression. In particular, the use of the current maintenance agents is associated with high rates of chronic kidney disease and metabolic and neoplastic complications. The search for improved treatment algorithms includes trial and error dose minimization or even complete withdrawal; most of which have been attempted with variable success rates. These strategies would be potentially more successful if more were known about long-term immune adaptive mechanisms that occur in response to transplantation. In addition, biomarkers signifying immunoregulation or tolerance, which could more clearly define the capability of withdrawing therapy, are needed to apply a more specific approach to patient management. This review will provide an overview of the mechanisms of liver transplant tolerance and the current and future strategies for immunosuppression withdrawal.

Regulatory T cell therapy: An option to induce operational tolerance in liver transplantation

Regulatory T cells (Treg) may play an important role in operational (clinical) tolerance (OT), a stable graft function without immunosuppression in an otherwise immunocompetent host, that is spontaneously observed in some patients many years after transplantation. Several ongoing clinical trials are currently testing the effects of donor-specific or non-specific Treg infusion with the goal to induce this state of OT a few months after liver transplantation (LT). The preliminary results of two of these trials have been recently published, and raise a number of comments and issues: (1) These two papers demonstrate that a 100 to 1000-fold ex-vivo expansion of Treg is possible in humans after 2 weeks of culture. The optimal human Treg dose is however not clearly established, and might be higher than the dose that would be expected from translating murine data. (2) A lot of concerns are remaining regarding the Treg purity before expansion, the Treg stability during in vitro culture and the in vivo fate of infused cells. A strict monitoring of Treg should thus be done at each step. (3) Since Treg may play a detrimental role in some conditions, such as viral diseases and cancer, potential LT recipients with such diseases should probably be excluded from the initial trials of Treg infusion. (4) The follow-up of tolerant liver recipients should include repeated liver biopsies and detection of autoantibodies and humoral response, in addition to conventional liver graft assessment, in order to prevent the development of immune complications related to immunosuppression withdrawal. (5) The final issue raised by Treg therapy in LT is the choice of the immunosuppressive regimen used before tapering or withdrawal, appropriate to preserve OT establishment.

NKT cells in the liver are important for periferal tolerance induction

Abstract NKT cells are mainly located in the liver in mice. The role of liver inherent NKT cells are still unclear. Method In this study, we employed mouse orthotopic liver transplantation and heterotopic heart transplantation models to critical examine the role of liver NKT cells in liver and peripheral tolerance induction. Results NKT cells were increased in the tolerated liver grafts which expressed higher levels of CD95L and PD-L1, while the CD4+CD25+Foxp3+ regulatory T cells (Treg) were markedly increased in the both liver and spleen at day 7 post transplantation. The heart allograft survival were prolonged significantly in the recipient which accepted donor spleen cells by portal vein (p.v.) injection in contract to the recipient which received the donor spleen cells by tail vein (i.v.) injection. In vitro immunological assay revealed that the number of NKT cells in the liver was increased and Treg were increased in both liver and spleen from the p.v. treated mice significantly. The IL-2 expression was decreased, IL-4 and IL-10 were increased. Further, significantly increased IL-4, IL-10, and IFN-γ production was detected from liver NPCs under aGalCer stimulation. Conclusion NKT cells are important to liver transplant tolerance induction, appear to play a key role in down regulation of peripheral immune responses and facilitate CD4+CD25+Foxp3+ Treg induction.

The mechanisms of Gamma delta T cells in the occurrence and development of hepatocellular carcinoma

Abstract γδ T cells are unique population of T cells in which their function still remain unclear. We have demonstrated that γδ T cell mainly located in the liver in mice. Our previous data based on liver transplantation model suggests that γδT cells are involved in the processes of liver transplant tolerance and rejection, and may play a role in the induction of liver graft rejection. In contrast to Foxp3+ CD4+ CD25+ Treg, the majority of γδT cells were mainly located in the liver and did not express TCRαβ and NK receptors, but expressed moderate levels of PD-1 and PD-L1 molecules. It has recently been shown that γδT cells play an important role against tumors and infectious diseases. Methods In this study we employed mouse orthotopic hepatocellular carcinoma model, critically examine the role of γδ T cells in the liver and peripheral on liver cancer development. Results The ratio of CD3+ T cells were decreased, but γδ T cells were increased in the surrounding tissue of the tumor and the spleen. Anti-γδ T cells antibody plus IL-2 treatment to tumor barrier mice could further increase the γδ T cells in the liver and spleen and limited the growth of the tumor. Conclusion γδ T cell may plays an important role in immunologic defence against hepatocellular carcinoma. More studies underlying the mechanisms and interplay among liver immune cells are under investigation.

Liver transplantation in the mouse: Insights into liver immunobiology, tissue injury, and allograft tolerance.

The surgically demanding mouse orthotopic liver transplant model was first described in 1991. It has proved to be a powerful research tool for the investigation of liver biology, tissue injury, the regulation of alloimmunity and tolerance induction, and the pathogenesis of specific liver diseases. Liver transplantation in mice has unique advantages over transplantation of the liver in larger species, such as the rat or pig, because the mouse genome is well characterized and there is much greater availability of both genetically modified animals and research reagents. Liver transplant experiments using various transgenic or gene knockout mice have provided valuable mechanistic insights into the immunobiology and pathobiology of the liver and the regulation of graft rejection and tolerance over the past 25 years. The molecular pathways identified in the regulation of tissue injury and promotion of liver transplant tolerance provide new potential targets for therapeutic intervention to control adverse inflammatory responses/immune-mediated events in the hepatic environment and systemically. In conclusion, orthotopic liver transplantation in the mouse is a valuable model for gaining improved insights into liver biology, immunopathology, and allograft tolerance that may result in therapeutic innovation in the liver and in the treatment of other diseases.

Tolerance in liver transplantation: Biomarkers and clinical relevance.

Transplantation is the optimal treatment for end-stage organ failure, and modern immunosuppression has allowed important progress in short-term outcomes. However, immunosuppression poorly influences chronic rejection and elicits chronic toxicity in current clinical practice. Thus, a major goal in transplantation is to understand and induce tolerance. It is well established that human regulatory T cells expressing the transcription factor FoxP3 play important roles in the maintenance of immunological self-tolerance and immune homeostasis. The major regulatory T cell subsets and mechanisms of expansion that are critical for induction and long-term maintenance of graft tolerance and survival are being actively investigated. Likewise, other immune cells, such as dendritic cells, monocyte/macrophages or natural killer cells, have been described as part of the process known as “operational tolerance”. However, translation of these results towards clinical practice needs solid tools to identify accurately and reliably patients who are going to be tolerant. In this way, a plethora of genetic and cellular biomarkers is raising and being validated worldwide in large multi-center clinical trials. Few of the studies performed so far have provided a detailed analysis of the impact of immunosuppression withdrawal on pre-existing complications derived from the long-term administration of immunosuppressive drugs and the side effects associated with them. The future of liver transplantation is aimed to develop new therapies which increase the actual low tolerant vs non-tolerant recipients ratio.

Role of NK, NKT cells and macrophages in liver transplantation.

Liver transplantation has become the treatment of choice for acute or chronic liver disease. Because the liver acts as an innate immunity-dominant organ, there are immunological differences between the liver and other organs. The specific features of hepatic natural killer (NK), NKT and Kupffer cells and their role in the mechanism of liver transplant rejection, tolerance and hepatic ischemia-reperfusion injury are discussed in this review.

Tolerance signatures in transplant recipients.

The intent of this review was to describe biomarkers that predict or identify individuals who exhibit tolerance to a transplanted organ. The identification of tolerance biomarkers would spare some individuals the toxicity of immunosuppressive agents, enhance the safety of studies to induce tolerance, and provide insights into mechanisms of tolerance that may aid in designing new regimens. Studies of tolerant kidney transplant recipients have revealed an association with B cells. More recent studies have suggested that these B cells may be less mature than from those in nontolerant recipients, and especially suited to suppress alloimmune responses. Biomarkers in tolerant liver transplant patients appear to be distinct from those associated renal tolerance. Most reports have identified an association with natural killer and/or γδ T cells rather than B cells. Recent data indicate biomarkers associated with iron homeostasis within the transplanted liver more accurately predict the tolerant state than do biomarkers expressed in the blood, suggesting that the renal allograft itself, which is infrequently sampled, would be informative. Given the encouraging progress in identifying tolerance biomarkers, it will be important to validate these markers in larger studies of transplant recipients undergoing prospective minimization or withdrawal of immunosuppression.

Rejection triggers liver transplant tolerance: Involvement of mesenchyme-mediated immune control mechanisms in mice.

Liver tolerance was initially recognized by the spontaneous acceptance of liver allografts in many species. The underlying mechanisms are not completely understood. However, liver transplant (LT) tolerance absolutely requires interferon (IFN)-γ, a rejection-associated inflammatory cytokine. In this study, we investigated the rejection of liver allografts deficient in the IFN-γ receptor and reveal that the liver graft is equipped with machineries capable of counterattacking the host immune response through a mesenchyme-mediated immune control (MMIC) mechanism. MMIC is triggered by T effector (Tef) cell-derived IFN-γ that drives expression of B7-H1 on graft mesenchymal cells leading to Tef cell apoptosis. We describe the negative feedback loop between graft mesenchymal and Tef cells that ultimately results in LT tolerance. Comparable elevations of T-regulatory cells and myeloid-derived suppressor cells were observed in both rejection and tolerance groups and were not dependent on IFN-γ stimulation, suggesting a critical role of Tef cell elimination in tolerance induction. We identify potent MMIC activity in hepatic stellate cells and liver sinusoidal endothelial cells. MMIC is unlikely exclusive to the liver, given that spontaneous acceptance of kidney allografts has been reported, although less commonly, probably reflecting variance in MMIC activity. MMIC may represent an important homeostatic mechanism that supports peripheral tolerance and could be a target for the prevention and treatment of transplant rejection. This study highlights that the graft is an active participant in the equipoise between tolerance and rejection and warrants more attention in the search for tolerance biomarkers.

Emergent Transcriptomic Technologies and Their Role in the Discovery of Biomarkers of Liver Transplant Tolerance.

Liver transplantation offers a unique window into transplant immunology due, in part, to the considerable proportion of recipients who develop immunological tolerance to their allograft. Biomarkers are able to identify and predict such a state of tolerance, and thereby able to establish suitable candidates for the minimization of hazardous immunosuppressive therapies, are not only of great potential clinical benefit but might also shed light on the immunological mechanisms underlying tolerance and rejection. Here, we review the emergent transcriptomic technologies serving as drivers of biomarker discovery, we appraise efforts to identify a molecular signature of liver allograft tolerance, and we consider the implications of this work on the mechanistic understanding of immunological tolerance.

Mesenchymal stromal cell-dependent reprogramming of Kupffer cells is mediated by TNF-α and PGE2 and is crucial for liver transplant tolerance.

The role of mesenchymal stromal cells (MSCs) in the modulation of liver transplant tolerance has attracted significant interest. However, the interaction between MSCs and Kupffer cells (KCs) has received little attention, and the effect of this interaction on liver transplant tolerance remains unclear. KCs were cultured in the presence and absence of MSCs. After 24 h, cells were treated with lipopolysaccharide (LPS), after which the production of cytokines and the expression of surface antigens were measured for cell function identification. Moreover, the effects of the KCs and the prostaglandin E2 (PGE2) levels produced by the MSCs were determined using an experimental rat liver transplantation model. Blood and liver samples were collected at three time points after transplantation for further analysis. After LPS treatment, when compared with the KC single cultures, the expression of pro-inflammatory cytokines (IL-1β, IL-6, MHC-II, CD40, CD80, and CD86) in the coculture system was down-regulated, whereas the expression of anti-inflammatory cytokines (TGF-β, IL-4, PGE2, and IL-10) was markedly increased. These data indicate that MSCs can reprogram the phenotype of KCs. However, KCs treated with miR/TNF-α (tumor necrosis factor) plasmid prior to coculture to inhibit the production of TNF-α resulted in an inhibition of the reprogramming effect of MSCs. Moreover, overexpression of PGE2 in MSCs increased the effect of MSCs on KC reprogramming. After rat liver transplantation, allograft recipients that received MSCs showed better allograft tolerance when compared with rats in which KC function was inhibited. Furthermore, rats treated with MSCs overexpressing PGE2 demonstrated the best liver tolerance of all of the groups tested. MSCs reprogram the phenotype of KCs through TNF-α and PGE2, and this process is crucial for the immunomodulatory function of MSCs in liver transplantation.

Galectin-1 in Stable Liver Transplant Recipients

IntroductionThe achievement of a state of tolerance and minimization of the immunosuppressive load form part of the “Holy Grail” in solid organ transplantation. Galectin-1 recently has been described to be involved in the maintenance of a tolerant environment, but there is no evidence of its role in human liver transplantation. The aim of our study was to measure the serum levels of galectin-1 in stable liver transplant recipients. MethodsSerum levels of galectin-1 were determined in 30 stable liver transplant recipients who had been free of rejection episodes for at least 8 years. Fifteen patients with an acute rejection episode and 34 healthy subjects were used as the control group. ResultsThe concentrations of galectin-1 were significantly higher in stable liver transplant recipients compared with healthy subjects and with the acute rejection group. ConclusionsThese preliminary results indicate that galectin-1 is upregulated in stable liver transplant recipients. Thus, our results extend the recent findings that galectin-1 may play an immune-suppressive role in liver transplantation. It remains to be established whether it might help to induce tolerance in liver transplantation.