Mouse Cardiac Allografts Research Articles

Stromal Cell-Derived Factor-1 Mediates Cardiac Allograft Tolerance Induced by Human Endometrial Regenerative Cell-Based Therapy.

Endometrial regenerative cells (ERCs) are mesenchymal‐like stromal cells, and their therapeutic potential has been tested in the prevention of renal ischemic reperfusion injury, acute liver injury, ulcerative colitis, and immunosuppression. However, their potential in the induction of transplant tolerance has not been investigated. The present study was undertaken to investigate the efficacy of ERCs in inducing cardiac allograft tolerance and the function of stromal cell‐derived factor‐1 (SDF‐1) in the ERC‐mediated immunoregulation. The inhibitory efficacy of human ERCs in the presence or absence of rapamycin was examined in both mouse cardiac allograft models between BALB/c (H‐2d) donors and C57BL/6 (H‐2b) recipients and in vitro cocultured splenocytes. AMD3100 was used to inhibit the function of SDF‐1. Intragraft antibody (IgG and IgM) deposition and immune cell (CD4+ and CD8+) infiltration were measured by immunohistochemical staining, and splenocyte phenotypes were determined by fluorescence‐activated cell sorting analysis. The results showed that ERC‐based therapy induced donor‐specific allograft tolerance, and functionally inhibiting SDF‐1 resulted in severe allograft rejection. The negative effects of inhibiting SDF‐1 on allograft survival were correlated with increased levels of intragraft antibodies and infiltrating immune cells, and also with reduced levels of regulatory immune cells including MHC class IIlowCD86lowCD40lowdendritic cells, CD68+CD206+macrophages, CD4+CD25+Foxp3+T cells, and CD1dhighCD5highCD83lowIL‐10highB cells both in vivo and in vitro. These data showed that human ERC‐based therapy induces cardiac allograft tolerance in mice, which is associated with SDF‐1 activity, suggesting that SDF‐1 mediates the immunosuppression of ERC‐based therapy for the induction of transplant tolerance. Stem Cells Translational Medicine 2017;6:1997–2008

Preoperative application of combination of portal venous injection of donor spleen cells and intraperitoneal injection of rapamycin prolongs the survival of cardiac allografts in mice.

To investigate the effects of preoperative portal venous injection of donor spleen cells (PVIDSC) and intraperitoneal injection of rapamycin in the acute rejection of cardiac allograft in mice and the underlying mechanisms. Homogenous female B6 mice and BALB/c mice were used as recipients and donors of heart transplantation. These mice were randomly divided into different groups and received PVIDSC alone, rapamycin alone, or PVIDSC and rapamycin combined therapy. In addition, the underlying mechanism was studied by measuring a number of cytokines. Preoperative combination of PVIDSC and intraperitoneal injection of rapamycin significantly prolonged the survival of heterotopic cardiac allograft in mice, but had no effects on the survival time of cardiac allografts in mice pre-sensitized by skin grafting. Preoperative combination of PVIDSC and intraperitoneal injection of rapamycin increased the expression of IL-10 and Foxp3 and reduced the expression of INF-. Short-term preoperative administration of rapamycin promotes the expression of CD4+CD25+Foxp3+ regulator T cells. However, preoperative using alone of rapamycin, or combination of PVIDSC and rapamycin had no effects on the inhibition of proliferation of memory T cells. Preoperative application of combination of PVIDSC and rapamycin significantly prolonged the survival time of cardiac allografts in mice but not in mice pre-sensitized by skin grafting. This may be explained by the fact that combination of PVIDSC and rapamycin inhibited the cellular immune response and induced the expression of IL-10 from Tr1 cells and CD4+CD25+FoxP3+ regulatory T cells.

Impact of hyperlipidemia on alloimmunity.

Hyperlipidemia is a comorbidity affecting a significant number of transplant patients despite treatment with cholesterol lowering drugs. Recently, it has been shown that hyperlipidemia can significantly alter T-cell responses to cardiac allografts in mice, and graft rejection is accelerated in dyslipidemic mice. Here, we review recent advances in our understanding of hyperlipidemia in graft rejection. Hyperlipidemic mice have significant increases in serum levels of proinflammatory cytokines, and neutralization of interleukin 17 (IL-17) slows graft rejection, suggesting that IL-17 production by Th17 cells was necessary but not sufficient for rejection. Hyperlipidemia also causes an increase in alloreactive T-cell responses prior to antigen exposure. Analysis of peripheral tolerance mechanisms indicated that this was at least in part due to alterations in FoxP3 T cells that led to reduced Treg function and the expansion of FoxP3 CD4 T cells expressing low levels of CD25. Functionally, alterations in Treg function prevented the ability to induce operational tolerance to fully allogeneic heart transplants through costimulatory-molecule blockade, a strategy that requires Tregs. These findings highlight the importance of considering the contribution of inflammatory comorbidities to cardiac allograft rejection, and point to the potential importance of managing hyperlipidemia in the transplant population.

Macrophage/monocyte-specific deletion of Ras homolog gene family member A (RhoA) downregulates fractalkine receptor and inhibits chronic rejection of mouse cardiac allografts

The cellular and molecular mechanisms of chronic rejection of transplanted organs remain obscure; however, macrophages are known to play a critical role in the injury and repair of allografts. Among multiple factors influencing macrophage infiltration to allografts, the fractalkine chemokine (C-X3-C motif) ligand 1(CX3CL1)/chemokine (C-X3-C motif) receptor 1 (CX3CR1) signaling pathway and actin cytoskeleton, which is regulated by a small guanosine-5׳-triphosphatase Ras homolog gene family member A (RhoA), are of the utmost importance. To define the role of macrophage/RhoA pathway involvement in chronic rejection, we generated mice with monocyte/macrophage-specific deletion of RhoA. Hearts from BALB/c (H-2d) donors were transplanted into RhoAflox/flox (no Cre) and heterozygous Lyz2Cre+/-RhoAflox/flox recipients treated with cytotoxic T-lymphocyte-associated protein 4 immunoglobulin to inhibit early T-cell response. Allografts were assessed for chronic rejection and monocyte/macrophage functions. The deletion of RhoA inhibited macrophage infiltration, neointimal hyperplasia of vasculature, and abrogated chronic rejection of the allografts. The RhoA deletion downregulated G protein-coupled fractalkine receptor CX3CR1, which activates the RhoA pathway and controls monocyte/macrophage trafficking into the vascular endothelium. This in turn promotes, through overproliferation and differentiation of smooth muscle cells in the arterial walls, neointimal hyperplasia. Our finding of codependence of chronic rejection on monocyte/macrophage CX3CR1/CX3CL1 and RhoA signaling pathways may lead to the development of novel anti-chronic rejection therapies.

Tracking of TCR-Transgenic T Cells Reveals That Multiple Mechanisms Maintain Cardiac Transplant Tolerance in Mice.

Solid organ transplantation tolerance can be achieved following select transient immunosuppressive regimens that result in long-lasting restraint of alloimmunity without affecting responses to other antigens. Transplantation tolerance has been observed in animal models following costimulation or coreceptor blockade therapies, and in a subset of patients through induction protocols that include donor bone marrow transplantation, or following withdrawal of immunosuppression. Previous data from our lab and others have shown that proinflammatory interventions that successfully prevent the induction of transplantation tolerance in mice often fail to break tolerance once it has been stably established. This suggests that established tolerance acquires resilience to proinflammatory insults, and prompted us to investigate the mechanisms that maintain a stable state of robust tolerance. Our results demonstrate that only a triple intervention of depleting CD25+ regulatory T cells (Tregs), blocking programmed death ligand-1 (PD-L1) signals, and transferring low numbers of alloreactive T cells was sufficient to break established tolerance. We infer from these observations that Tregs and PD-1/PD-L1 signals cooperate to preserve a low alloreactive T cell frequency to maintain tolerance. Thus, therapeutic protocols designed to induce multiple parallel mechanisms of peripheral tolerance may be necessary to achieve robust transplantation tolerance capable of maintaining one allograft for life in the clinic.

Platelet-derived Growth Factor-B Protects Rat Cardiac Allografts From Ischemia-reperfusion Injury.

Microvascular dysfunction and cardiomyocyte injury are hallmarks of ischemia-reperfusion injury (IRI) after heart transplantation. Platelet-derived growth factors (PDGF) have an ambiguous role in this deleterious cascade. On one hand, PDGF may exert vascular stabilizing and antiapoptotic actions through endothelial-pericyte and endothelial-cardiomyocyte crosstalk in the heart; and on the other hand, PDGF signaling mediates neointimal formation and exacerbates chronic rejection in cardiac allografts. The balance between these potentially harmful and beneficial actions determines the final outcome of cardiac allografts. We transplanted cardiac allografts from Dark Agouti rat and Balb mouse donors to fully major histocompatibility complex-mismatched Wistar Furth rat or C57 mouse recipients with a clinically relevant 2-hour cold ischemia and 1-hour warm ischemia. Ex vivo intracoronary delivery of adenovirus-mediated gene transfer of recombinant human PDGF-BB upregulated messenger RNA expression of anti-mesenchymal transition and survival factors BMP-7 and Bcl-2 and preserved capillary density in rat cardiac allografts at day 10. In mouse cardiac allografts PDGF receptor-β, but not -α intragraft messenger RNA levels were reduced and capillary protein localization was lost during IRI. The PDGF receptor tyrosine kinase inhibitor imatinib mesylate and a monoclonal antibody against PDGF receptor-α enhanced myocardial damage evidenced by serum cardiac troponin T release in the rat and mouse cardiac allografts 6 hours after reperfusion, respectively. Moreover, imatinib mesylate enhanced rat cardiac allograft vasculopathy, cardiac fibrosis, and late allograft loss at day 56. Our results suggest that PDGF-B signaling may play a role in endothelial and cardiomyocyte recovery from IRI after heart transplantation.

Molecular Chimeric Recipient Precursor T Cells Promote Cardiac Allograft Survival in Mice

BackgroundMolecular chimerism has become a potential method to induce donor-specific transplant tolerance. We researched the prolongation of cardiac allograft survival by recipient mouse molecular chimeric precursor T cells (pre-T cells) or hematopoietic stem cells (HSCs) infusion in vivo. MethodsThe donor C57BL/6 mouse MHC-I gene (H-2Kb and H-2Db gene) were amplified by RT-PCR. The identified recipient BALB/c mouse pre-T cells and HSCs were transduced by the pMSCVneo retroviral vector of C57BL/6 mouse MHC-I gene (pMSCVneo-H-2Db/H-2Kb). Then the molecular chimeric cells were transfused back to the BALB/c mice. Allogeneic T-lymphocyte proliferation was assessed in mixed lymphocyte reactions (MLR). A mouse model of heterotopic abdominal heart transplantation was performed to evaluate survival times and histological grade of acute rejection at 7 days after transplantation. ResultsBALB/c mice molecular chimeric pre-T cells and HSCs were cultured successfully after pMSCV-H-2Db/H-2Kb transduction. After the molecular chimeric pre-T cell treatment, the result of MLR showed that the stimulating index of allogeneic T lymphocyte had a statistically significant decrease, which also exhibited a significant reduction after molecular chimeric HSC treatment. The survival time of cardiac allograft was prolonged after chimeric pre-T cell or HSC infusion; meanwhile, pathologic rejection grade decreased significantly. Nevertheless, molecular chimeric pre-T cells exhibited a longer median survival time. ConclusionThe molecular chimeric recipient mouse pre-T cell or HSC infusion reduced spleen T cells' response to allogeneic T cells in vitro and delayed cardiac allograft rejection in vivo. Pre-T cells have more advantages than HSCs on the prolongation of mouse cardiac allograft survival.

Hyperlipidemia Alters Regulatory T Cell Function and Promotes Resistance to Tolerance Induction Through Costimulatory Molecule Blockade

Recent work from our laboratory has shown that hyperlipidemia promotes accelerated rejection of vascularized cardiac allografts in mice by inducing anti-donor Th17 reactivity and production of IL-17. Here, we show that hyperlipidemia also affects FoxP3(+) regulatory T cells (Tregs). Hyperlipidemia promotes the development of Tregs that express low levels of CD25. Hyperlipidemia also promotes a decrease in central Tregs and an increase in effector Tregs that appears to account for the increase in the frequency of CD25(low) Tregs. Alterations in Treg subsets also appear to lead to alterations in Treg function. The ability of FoxP3(+) , CD25(high) , CD4(+) Tregs from hyperlipidemic mice to inhibit proliferation of effector T cells stimulated with anti-CD3 and CD28 was reduced when compared with Tregs from control mice. Regulatory T cells isolated from hyperlipidemic recipients exhibit increased activation of Akt, and a reduction in Bim levels that permits the expansion of FoxP3(+) CD25(low) CD4(+) T cells. Hyperlipidemic mice were also resistant to tolerance induction using costimulatory molecule blockade consisting of anti-CD154 and CTLA4Ig, a strategy that requires Tregs. Together, our data suggest that hyperlipidemia profoundly affects Treg subsets and function as well as the ability to induce tolerance.

Galectin-9 in Combination With EX-527 Prolongs the Survival of Cardiac Allografts in Mice After Cardiac Transplantation

Galectin-9 (Gal-9), a member of the galectin family, has a variety of biologic activities. However, its role in allografts is not fully clarified yet. The relationship between interleukin-17 (IL-17) and Gal-9 and the role of Gal-9 in TH17-cell differentiation also remain unclear. We built a murine cardiac transplantation model, which we treated with Gal-9 and/or EX-527, a specific Sirtuin-1 inhibitor. Afterwards, flow-cytometric analysis and reverse-transcription polymerase chain reaction were used to detect the number of TH17 cells and the expression of key factors involved in the differentiation of TH17 cells; in addition, the survival times of cardiac transplanted mice in different groups were recorded. The levels of circulating TH17 cells were found to increase in the peripheral blood of mice that exhibited acute rejection (AR) after heart transplantation, which was determined to be correlated with the rejection grade. Gal-9 or EX-527 can inhibit the activation and differentiation of TH17 cells and effectively suppress TH17-cell–mediated AR. These data provide new evidence for the potential regulatory effects of Gal-9 in alloimmune responses in a murine model of heart transplantation, and suggest the combined use of galectin-9 and EX-527 may be a powerful method to induce tolerance of fully mismatched murine cardiac allografts.

Permanent acceptance of mouse cardiac allografts with CD40 siRNA to induce regulatory myeloid cells by use of a novel polysaccharide siRNA delivery system.

The CD40/CD154 co-stimulatory pathway is crucial in alloimmune response. We developed a novel small interfering RNA (siRNA) delivery system with a poly-dA extension at the 5'-end of the siRNA sense strand that was stably incorporated into 1,3-β-glucan (schizophyllan, SPG). This was captured and incorporated into dendritic cells (DCs) through its receptor, Dectin-1, specifically silencing CD40 genes (siCD40) to exert immunoregulatory activity. siCD40/SPG-treated CBA mice permanently accepted B10 fully mismatched cardiac allografts. Consistent with graft survival, the infiltration of CD4(+), CD8(+) T cells into the graft was lower, and that the numbers of CD40(low)CD11c(+) DCs cells and CD4(+)Foxp3(+)cells were increased in both the graft and in the recipient spleen. In addition, naive CBA recipients given an adoptive transfer of splenocytes from the primary recipients with siCD40/SPG accepted a heart graft from donor-type B10, but not third-party Balb/c mice. In conclusion, the treatment with siCD40/SPG targeting DCs could generate antigen-specific Tregs, resulting in the permanent acceptance of mouse cardiac allografts. These findings have important implications for clarifying the mechanism underlying the induction of tolerance in DCs, and also highlight the potential of immunomodulation and the feasibility of siRNA-based clinical therapy in the transplantation field.

CD4(+)CD25 (+) regulatory T cells are not required for mesenchymal stem cell function in fully MHC-mismatched mouse cardiac transplantation.

Although the immunomodulative properties of mesenchymal stem cells (MSCs) open up attractive possibilities in solid-organ transplantation, information concerning the optimal dose, route, timing of administration, major histocompatibility complex (MHC)-restriction and relevant mechanisms is currently lacking. Therefore, better characterization of MSC immunoregulatory activity and elucidation of its mechanisms are crucial. In this study, we confirmed that MSCs did not elicit proliferation by allogeneic CD4(+) T cells, suggesting that MSCs were not immunogenic. By using C57BL/6 mouse MSCs as donor-derived or recipient-derived or as third-party MSCs, we discovered that MSCs suppressed CD4(+) T cell proliferation and prolonged mouse cardiac allograft survival in a dose-dependent and non-MHC-restricted manner. We also found that intraperitoneal administration favored survival prolongation, although this prolongation was weaker than that via the intravenous route. Only infusion at earlier time points favored survival prolongation. Depletion of CD4(+)CD25(+) T cells did not affect the immunosuppression of MSCs on CD4(+) T cells. Moreover, MSCs did not induce regulatory T cells. The in vivo data revealed that MSCs did not increase the percentage of CD4(+)CD25(+) T cells and FoxP3 expression. More importantly, we demonstrated for the first time that depletion of CD4(+)CD25(+) T cells did not hinder MSC-induced survival prolongation, indicating that CD4(+)CD25(+) regulatory T cells were not essential for the prolongation of MSC-mediated allograft survival.

Requirement of B7-H1 in mesenchymal stem cells for immune tolerance to cardiac allografts in combination therapy with rapamycin

BackgroundThe potential of mesenchymal stem cells (MSCs) for immunosuppression has been tested in transplantation, but its mechanisms are not fully understood. This study investigated the role of MSC-expressing B7-H1 in the induction of immune tolerance to cardiac allografts by the combination therapy of MSCs and rapamycin (RAPA). MethodsThe anti-alloimmunity of donor MSCs in the presence or absence of RAPA was examined in both mouse cardiac allograft model (C57BL/6 to BALB/c mice) and a variety of cultured immune cells. Immunohistochemical staining was used for the measurement of intragraft antibody deposition, and fluorescence-activated cell sorting (FACS) for the determination of serum alloantibodies and leukocyte phenotypes. ResultsB7-H1 expression in cultured MSCs was up-regulated following IFN-γ stimulation. In transplant recipients, combination therapy of MSCs and RAPA induced immune tolerance to allografts, but blockade of B7-H1 on MSCs with monoclonal antibody abrogated the combination therapy-induced immune tolerance as heart allografts were rejected. The negative effect of MSC-expressing B7-H1 neutralization on graft survival was correlated with a reduction of regulatory immune cells (CD4+CD25+Foxp3+ T cells, tolerogenic dendritic cells and IL-4highIL-10HighCD83low B cells), and also with an increase in alloantibody (IgG and IgM) levels both inside the grafts and in the circulation as compared with un-neutralized controls. In vitro MSC-mediated suppression of antibody production and B cell proliferation depended on B7-H1 function and cell contact between CD19+ B cells and MSCs. ConclusionThese data suggest that MSC-expressing B7-H1 mediates the immune tolerance to cardiac allografts in recipients receiving MSC and RAPA combination therapy.

(435) - Donor Single-dose Treatment with VEGFR-3 Antibody Reduces Acute Alloimmune Response by Targeting Lymphatic Endothelial Cell Activation

VEGF-C - VEGFR-3 signaling may regulate the activation of lymphatic endothelial cells and the initiation of alloimmunity after transplantation. We investigated the impact of cold ischemic preservation and ischemia-reperfusion (I/R) injury on the activation of lymphatic endothelial cells and evaluated the effect of donor treatment with VEGFR-3 antibody on subsequent development of acute alloimmune response in rat and mouse cardiac allografts.

Endogenous Memory CD8 T Cells Are Activated Within Cardiac Allografts Without Mediating Rejection

Endogenous memory CD8 T cells infiltrate MHC-mismatched cardiac allografts within 12–24 h posttransplant in mice and are activated to proliferate and produce IFN-γ. To more accurately assess the graft injury directly imposed by these endogenous memory CD8 T cells, we took advantage of the ability of anti-LFA-1 mAb given to allograft recipients on days 3 and 4 posttransplant to inhibit the generation of primary effector T cells. When compared to grafts from IgG-treated recipients on day 7 posttransplant, allografts from anti-LFA-1 mAb-treated recipients had increased numbers of CD8 T cells but these grafts had marked decreases in expression levels of mRNA encoding effector mediators associated with graft injury and decreases in donor-reactive CD8 T cells producing IFN-γ. Despite this decreased activity within the allograft, CD8 T cells in allografts from recipients treated with anti-LFA-1 mAb continued to proliferate up to day 7 posttransplant and did not upregulate expression of the exhaustion marker LAG-3 but did have decreased expression of ICOS. These results indicate that endogenous memory CD8 T cells infiltrate and proliferate in cardiac allografts in mice but do not express sufficient levels of functions to mediate overt graft injury and acute rejection.

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

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

Galectin-7 in Cardiac Allografts in Mice: Increased Expression Compared With Isografts and Localization in Infiltrating Lymphocytes and Vascular Endothelial Cells

We sought to identify elevated expression of galectin-7, a T-cell–binding protein, in renal allograft recipients undergoing acute rejection episodes. Allografts and isografts were examined immunohistologically and using quantitative Western blot analysis for galectin-7 protein. The expression of galectin-7 in T lymphocytes from draining lymph nodes in the recipient mice was examined using flow cytometry. We observed galectin-7 to gradually increase with time in the allografts to be significantly higher than that in isografts at days 3, 5, and 7 postoperative: 0.85 ± 0.03 versus 0.69 ± 0.05; 1.15 ± 0.11 versus 0.81 ± 0.02; and 2.02 ± 0.12 versus 0.81 ± 0.05 (P < .05). The majority of galectin-7 was located in the infiltrating lymphocytes and vascular endothelial cells. Furthermore, the percentage of CD4+galectin-7+ and CD8+galectin-7+ T cells from draining lymph nodes in the allograft group was higher than that in the isograft group: 28.0 ± 1.0% versus 1.2 ± 0.2%; and 12.4 ± 0.8% versus 0.4 ± 0.1% (P < .01). In conclusion, galectin-7 relates to acute allograft rejection and T-cell responses possibly as an accelerant.

A Novel Sirna Delivery System Specifically Silencing Target Gene to Induce Permanent Acceptance of Mouse Cardiac Allograft

A Novel Sirna Delivery System Specifically Silencing Target Gene to Induce Permanent Acceptance of Mouse Cardiac Allograft

5-Aminolaevulinic Acid (ALA) Induces Permanent Acceptance of Mouse Cardiac Allograft by Increasing Regulatory T Cells

5-Aminolaevulinic Acid (ALA) Induces Permanent Acceptance of Mouse Cardiac Allograft by Increasing Regulatory T Cells

Tolerance Induction by Exosomes from Immature Dendritic Cells and Rapamycin in a Mouse Cardiac Allograft Model

BackgroundDendritic cells (DCs) release bioactive exosomes that play an important role in immune regulation. Because they express low levels of class I major histocompatibility complex (MHC) and co-stimulatory molecules, exosomes derived from donor immature DCs (imDex) prolong allograft survival by inhibiting T-cell activation. However, this effect is limited and does not induce immunological tolerance when imDex are administered alone. Thus, we tested the effect of combined treatment with donor imDex and low-dose rapamycin on inducing tolerance in a mouse cardiac transplantation model.MethodsImDex were obtained from the culture supernatant of immature DCs derived from donor mouse (C57BL/6) bone marrow and were injected with suboptimal doses of rapamycin into recipient mouse (BALB/c) before and after transplantation. The capacity of this treatment to induce immune tolerance was analyzed in vitro and in vivo using the mouse cardiac transplantation model.ResultsDonor imDex expressed moderate levels of MHC class II and low levels of MHC class I and co-stimulatory molecules, but neither imDex nor subtherapeutic rapamycin dose alone induced cardiac allograft tolerance. Combined treatment with imDex and rapamycin, however, led to donor specific cardiac allograft tolerance. This effect was accompanied by decreased anti-donor antigen cellular response and an increased percentage of spleen CD4+CD25+ T cells in recipients. Furthermore, this donor specific tolerance could be further transferred to naïve allograft recipients through injection of splenocytes, but not serum, from tolerant recipients.ConclusionCombined with immunosuppressive treatment, donor imDex can prolong cardiac allograft survival and induce donor specific allograft tolerance.

Gene Transfer of Heme Oxygenase-1 Using an Adeno-Associated Virus Serotype 6 Vector Prolongs Cardiac Allograft Survival

Introduction. Allograft survival can be prolonged by overexpression of cytoprotective genes such as heme oxygenase-1 (HO-1). Modifications in vector design and delivery have provided new opportunities to safely and effectively administer HO-1 into the heart prior to transplantation to improve long-term graft outcome. Methods. HO-1 was delivered to the donor heart using an adeno-associated virus vector (AAV) with a pseudotype 6 capsid and vascular endothelial growth factor (VEGF) to enhance myocardial tropism and microvascular permeability. Survival of mouse cardiac allografts, fully or partially mismatched at the MHC, was determined with and without cyclosporine A. Intragraft cytokine gene expression was examined by PCR. Results. The use of AAV6 to deliver HO-1 to the donor heart, combined with immunosuppression, prolonged allograft survival by 55.3% when donor and recipient were completely mismatched at the MHC and by 94.6% if partially mismatched. The combination of gene therapy and immunosuppression was more beneficial than treatment with either AAV6-HO-1 or CsA alone. IL-17a, b, e and f were induced in the heart at rejection. Conclusions. Pretreatment of cardiac allografts with AAV6-HO-1 plus cyclosporine A prolonged graft survival. HO-1 gene therapy represents a beneficial adjunct to immunosuppressive therapy in cardiac transplantation.