Biomedical subjects

M H Sayegh

Publications and source records attributed to M H Sayegh.

At least 19 recordsLinked to original sources

PubMed2001

Recipient MHC class II expression is required to achieve long-term survival of murine cardiac allografts after costimulatory blockade.

To study the role of the direct and indirect pathways in achieving tolerance, we used genetically altered mouse strains in two ways: 1) MHC class II-deficient mice were used as donors of skin and cardiac grafts to eliminate the direct CD4(+) T cell response, and 2) B6 II(-)4(+) mice, which are MHC class II-deficient mice expressing an MHC class II transgene only on thymic epithelium, were used as recipients of normal grafts. These mice cannot mount an indirect response. Eliminating the indirect pathway actually made it more difficult to achieve prolonged allograft survival when we used costimulatory blockade than when both pathways were available. Costimulatory blockade was ineffective even when CD4(+) T cells from normal animals were transferred into recipients that lacked MHC class II molecules. These results suggest that an active CD4(+) response through the indirect pathway is necessary for costimulatory blockade to be effective in prolonging allograft survival.

Abatacept↗

PubMed2001

Rejection of mouse cardiac allografts by costimulation in trans.

The activation of T cells by B7 costimulation in trans has been demonstrated in vitro, but the in vivo relevance is unknown. To study costimulation in trans of CD4(+) T cells in vivo, we performed cardiac transplants from B7-1/B7-2-deficient mice to recipients that do not express MHC class II molecules on peripheral APCs, but do have functional CD4(+) T cells (II(-)/4(+) mice). This model restricts the B7-dependent activation of CD4(+) T cells to costimulation in trans and excludes any contribution from indirect Ag presentation. We find that II(-)/4(+) recipients reject B7-deficient grafts as rapidly as wild-type grafts, suggesting that costimulation in trans can mediate rejection as potently as costimulation in cis. Treatment of II(-)/4(+) recipients of B7-deficient grafts with depleting Abs to CD4 or CD8 demonstrates that indirect Ag presentation to CD8(+) cells does not significantly contribute to rejection. This is the first demonstration that costimulation in trans can mediate an immune response in vivo and has important therapeutic implications.

Animals↗

PubMed2001

CD28-independent costimulation of T cells in alloimmune responses.

T cell costimulation by B7 molecules plays an important role in the regulation of alloimmune responses. Although both B7-1 and B7-2 bind CD28 and CTLA-4 on T cells, the role of B7-1 and B7-2 signaling through CTLA-4 in regulating alloimmune responses is incompletely understood. To address this question, we transplanted CD28-deficient mice with fully allogeneic vascularized cardiac allografts and studied the effect of selective blockade of B7-1 or B7-2. These mice reject their grafts by a mechanism that involves both CD4(+) and CD8(+) T cells. Blockade of CTLA-4 or B7-1 significantly accelerated graft rejection. In contrast, B7-2 blockade significantly prolonged allograft survival and, unexpectedly, reversed the acceleration of graft rejection caused by CTLA-4 blockade. Furthermore, B7-2 blockade prolonged graft survival in recipients that were both CD28 and CTLA-4 deficient. Our data indicate that B7-1 is the dominant ligand for CTLA-4-mediated down-regulation of alloimmune responses in vivo and suggest that B7-2 has an additional receptor other than CD28 and CTLA-4 to provide a positive costimulatory signal for T cells.

Abatacept↗

PubMed2001

Involvement of the direct and indirect pathways of allorecognition in tolerance induction.

It is generally accepted that there are two pathways of allorecognition, direct and indirect, that together contribute to allograft rejection. Although it has been suggested that the direct pathway predominates during early acute rejection and that the indirect pathway provides a continuous supply of alloantigen responsible for chronic rejection, the true relative contribution of each pathway to the overall rejection process is still not entirely known. It is clear, however, that any strategies designed to achieve the ultimate goal in transplantation, the induction of tolerance, will need to take into account both pathways. This review seeks to explore the involvement of the direct and indirect pathways of allorecognition on a mechanistic level as it relates to the induction of tolerance. A brief historical perspective is included for each pathway as well as a comprehensive review of the mechanisms felt to be active during tolerance induction.

Animals↗

PubMed2001

Tolerance and chronic rejection.

The most common cause of chronic allograft loss is an incompletely understood clinicopathological entity called chronic rejection (CR). Recent reports suggest an improvement in long-term renal allograft survival, although it is not clear from these data whether a true reduction of biopsy-proven CR has occurred. Although newer immunosuppressive medications have greatly reduced the incidence of acute rejection (AR) in the early post-transplantation period, the ideal therapy for both AR and CR would be to achieve a state of tolerance. By definition, such a state should allow for indefinite allograft survival, with no histopathological evidence of CR, despite immunocompetence in the host (i.e. without the need for chronic immunosuppression). Although several experimental studies are able to achieve tolerance, with clear improvement in allograft survival, detailed studies on graft function and morphology are often not included. This review will discuss possible ways that tolerance induction could lead to a CR-free state. General mechanisms of CR and transplantation tolerance induction are discussed as well as the difficulties in translating small animals studies into large animals and humans.

Animals↗

PubMed2001

Indirect recognition of allopeptides promotes the development of cardiac allograft vasculopathy.

Graft loss from chronic rejection has become the major obstacle to the long-term success of whole organ transplantation. In cardiac allografts, chronic rejection is manifested as a diffuse and accelerated form of arteriosclerosis, termed cardiac allograft vasculopathy. It has been suggested that T-cell recognition of processed alloantigens (allopeptides) presented by recipient antigen-presenting cells through the indirect pathway of allorecognition plays a critical role in the development and progression of chronic rejection. However, definitive preclinical evidence to support this hypothesis is lacking. To examine the role of indirect allorecognition in a clinically relevant large animal model of cardiac allograft vasculopathy, we immunized MHC inbred miniature swine with synthetic polymorphic peptides spanning the alpha(1) domain of an allogeneic donor-derived swine leukocyte antigen class I gene. Pigs immunized with swine leukocyte antigen class I allopeptides showed in vitro proliferative responses and in vivo delayed-type hypersensitivity responses to the allogeneic peptides. Donor MHC class I disparate hearts transplanted into peptide-immunized cyclosporine-treated pigs not only rejected faster than unimmunized cyclosporine-treated controls (mean survival time = 5.5 +/-1.7 vs. 54.7 +/-3.8 days, P < 0.001), but they also developed obstructive fibroproliferative coronary artery lesions much earlier than unimmunized controls (<9 vs. >30 days). These results definitively link indirect allorecognition and cardiac allograft vasculopathy.

Animals↗

PubMed2001

Specific MDR1 P-glycoprotein blockade inhibits human alloimmune T cell activation in vitro.

MDR1 P-glycoprotein (P-gp), the multidrug resistance-associated transmembrane transporter, is physiologically expressed by human peripheral immune cells, but its role in cell-mediated immunity remains poorly understood. Here, we demonstrate a novel role for P-gp in alloantigen-dependent human T cell activation. The pharmacologic P-gp inhibitor tamoxifen (1-10 microM) and the MDR1 P-gp-specific mAb Hyb-241 (1-20 microg/ml), which detected surface P-gp on 21% of human CD3(+) T cells and 84% of CD14(+) APCs in our studies, inhibited alloantigen-dependent, but not mitogen-dependent, T cell proliferation in a dose-dependent manner from 40-90% (p < 0.01). The specific inhibitory effect on alloimmune T cell activation was associated with >85% inhibition (p < 0.01) of IL-2, IFN-gamma, and TNF-alpha production in 48-h MLR coculture supernatants. Addition of recombinant human IL-2 (0.1-10 ng/ml) restored proliferation in tamoxifen-treated cocultures. Pretreatment of purified CD4(+) T cells with Hyb-241 mAb before coculture resulted in inhibition of CD4(+) T cellular IFN-gamma secretion. Also, blockade of P-gp on allogeneic APCs inhibited IL-12 secretion. Taken together these results demonstrate that P-gp is functional on both CD4(+) T cells and CD14(+) APCs, and that P-gp blockade may attenuate both IFN-gamma and IL-12 through a positive feedback loop. Our results define a novel role for P-gp in alloimmunity and thus raise the intriguing possibility that P-gp may represent a novel therapeutic target in allograft rejection.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed2001

CTLA4Ig-induced linked regulation of allogeneic T cell responses.

The mechanisms by which CTLA4Ig exerts its powerful immunomodulatory effects are not clear. We show here that CTLA4Ig can induce linked regulation of allogeneic porcine T cell responses in vitro. Naive miniature swine SLA(dd) T cells were rendered hyporesponsive to specific allogeneic Ag after coculturing with MHC-mismatched SLA(cc) stimulators in the presence of CTLA4Ig. These Ag-specific hyporesponsive T cells were subsequently able to actively inhibit the allogeneic responses of naive syngeneic T cells in an MHC-linked fashion, as the responses of naive SLA(dd) responders against specific SLA(cc) and (SLA(ac))F(1) stimulators were inhibited, but allogeneic responses against a 1:1 mixture of SLA(aa) (I(a), II(a)) and SLA(cc) (I(c), II(c)) were maintained. This inhibition could be generated against either class I or class II Ags, was radiosensitive, and required cell-cell contact. Furthermore, the mechanism of inhibition was not dependent upon a deletional, apoptotic pathway, but it was reversed by anti-IL-10 mAb. These data suggest that CTLA4Ig-induced inhibition of naive allogeneic T cell responses can be mediated through the generation of regulatory T cells via an IL-10-dependent mechanism.

Abatacept↗

PubMed2001

Quantifying the frequency of alloreactive T cells in vivo: new answers to an old question.

Alloreactive T cell precursor frequency was measured in vivo using fluorescent dye labeling in combination with novel models based on lymphocyte activation and recovery. CFSE-labeled C57BL/6 (H-2(b)) spleen and lymph node cells were adoptively transferred to C57BL/6xDBA F(1) (H-2(b/d)) recipients, a parent-->F(1) MHC mismatch in which only donor cells respond. Recipients were sacrificed at serial time points to assess engraftment efficiency, and the extent of donor cell activation and proliferation. These data were used to calculate alloreactive T cell frequencies that varied 30-fold (0.71 +/- 0.31% to 21.05 +/- 3.62%), depending upon whether it was assumed that all donor cells injected became established and were capable of responding, or that only those present at later time points (24-72 h) were available to respond. By measuring the number of cells established in the recipient 24 h after transfer, before proliferation, we calculated an in vivo alloreactive frequency of approximately 7%. Using CD69 expression at 48 h to quantify activation, we found that 40-50% of the alloactivated CD4(+) donor T cells do not divide. Studies of cotransferred congenic and allogeneic cells demonstrated that bystander proliferation does not occur. We conclude that accurate calculations of alloreactive precursor frequency must account for both proliferation and cell engraftment. When this is done, a high percentage of alloreactive T cells exists across an MHC mismatch, but not all alloreactive cells proliferate in vivo. Bystander proliferation is negligible, revealing exquisite specificity to the alloresponse. These data provide a novel approach to quantify alloreactive T cell responses during specific immunomodulatory strategies in vivo.

Adoptive Transfer↗

PubMed2001

Targeting signal 1 through CD45RB synergizes with CD40 ligand blockade and promotes long term engraftment and tolerance in stringent transplant models.

The induction and maintenance of allograft tolerance is a daunting challenge. Although combined blockade of CD28 and CD40 ligand (CD40L)-costimulatory pathways prevents allograft rejection in some murine models, this strategy is unable to sustain engraftment in the most immunogenic allograft and strain combinations. By targeting T cell activation signals 1 and 2 with the novel combination of anti-CD45RB and anti-CD40L, we now demonstrate potent enhancement of engraftment in C57BL/6 recipients that are relatively resistant to costimulatory blockade. This combination significantly augments the induction of tolerance to islet allografts and dramatically prolongs primary skin allograft survival. Compared with either agent alone, anti-CD45RB plus anti-CD40L inhibits periislet infiltration by CD8 cells, B cells, and monocytes; inhibits Th1 cytokines; and increases Th2 cytokine expression within the graft. These data indicate that interference with activation signals one and two may provide synergy essential for prolonged engraftment in situations where costimulatory blockade is only partially effective.

Adjuvants, Immunologic↗

PubMed2001

CD28-B7-mediated T cell costimulation in chronic cardiac allograft rejection: differential role of B7-1 in initiation versus progression of graft arteriosclerosis.

Provision of adequate T cell costimulation is critical for the development of acute and chronic allograft rejection. We have previously reported that early blockade of CD28-B7 T cell costimulation prevents the development of graft arteriosclerosis, in the LEW into F344 rat cardiac transplant model. In this study, we used the same model to examine the requirement for CD28-B7-mediated T cell costimulation in the progression of established chronic rejection and examined the individual roles of B7-1 (CD80) and B7-2 (CD86) costimulatory molecules. Late blockade of CD28-B7 T cell costimulation by the fusion protein CTLA4Ig, which binds both CD80 and CD86, attenuated the development of transplant arteriosclerosis, mononuclear cell infiltration, and parenchymal fibrosis in this model. Selective blockade of CD80 using the mutant fusion protein Y100F was as effective as CTLA4Ig in this regard. In contrast to CTLA4Ig, blockade of CD80 alone by Y100F was ineffective at preventing early graft loss and prolonging graft survival when given early after transplantation. This study is the first to demonstrate that late blockade of CD28-B7 T cell costimulation interrupts chronic cardiac allograft rejection, and it indicates the importance of continued T cell activation in this process. This study further defines functional differences between CD80 and CD86 costimulatory molecules in vivo.

Abatacept↗

PubMed2001

Indirect allorecognition of mismatched donor HLA class II peptides in lung transplant recipients with bronchiolitis obliterans syndrome.

A correlation between indirect allorecognition of mismatched donor HLA class I peptides and development of bronchiolitis obliterans syndrome (BOS) after lung transplantation has been previously observed. The aim of this study was to determine whether there was a correlation between indirect allorecognition of mismatched donor HLA class II peptides and development of BOS after lung transplantation. Peripheral blood mononuclear cells from nine BOS+ and nine BOS-lung transplant recipients were cultured with synthetic peptides corresponding to the beta-chain hypervariable region of a mismatched donor HLA-DR molecule. Then, proliferative alloreactivity as well as frequency of alloreactive T cells were determined. In addition, the immunodominant epitopes from the donor HLA-DR molecules were identified in selected patients. T cells from BOS+ patients showed a dose-dependent proliferative alloreactivity against donor HLA-DR peptides that was significantly higher than that observed in BOS- patients (p=0.001). Similarly, the frequency of HLA-DR alloreactive T cells was significantly higher in BOS+ patients than in BOS- patients (p=0.001). This T-cell alloreactivity was directed against a single immunodominant HLA-DR peptide. These results suggest that indirect alloreactivity to donor HLA class II molecules may play a role in the pathogenesis of BOS after lung transplantation.

Amino Acid Sequence↗

PubMed2001

Membranous lupus nephritis in a renal allograft: response to mycophenolate mofetil therapy.

Membranous lupus nephritis in a renal allograft is considered rare. A 43-year-old man with quiescent systemic lupus erythematosus (SLE) received a HLA identical transplant from his sister and 4 years later developed persistent nephrotic range proteinuria and morphological features most compatible with membranous lupus nephritis on biopsy. Angiotensin-converting enzyme (ACE) inhibitors and angiotensin II receptor antagonists, although successful in reducing proteinuria, were associated on three occasions with acute allograft dysfunction. Sustained reduction of proteinuria and stable graft function were achieved using mycophenolate mofetil (MMF). MMF is emerging as a new therapy for primary renal disease in SLE. This is the first report of successful treatment of membranous lupus nephritis in an allograft using MMF. We review all cases of transplant-associated membranous lupus nephritis in the English literature.

Adult↗

PubMed2001

CTLA-4 up-regulation plays a role in tolerance mediated by CD45.

Cytolytic T lymphocyte-associated antigen 4 (CTLA-4) is a critical down-regulatory molecule in T cells that plays a major role in peripheral tolerance. Although the CD45 protein tyrosine phosphatase is a potent immunomodulatory target, the mechanisms by which antibody against CD45RB isoforms (anti-CD45RB) induces allograft tolerance remain unclear. We show here that anti-CD45RB treatment alters CD45 isoform expression on T cells, which is associated with rapid up-regulation of CTLA-4 expression. These effects appear specific and occur without up-regulation of other activation markers. Administration of a blocking monoclonal antibody to CTLA-4 at the time of transplantation prevents anti-CD45RB therapy from prolonging islet allograft survival. In addition, treatment with cyclosporin A blocks anti-CD45RB-induced CTLA-4 expression and promotes acute rejection. These data suggest that anti-CD45RB acts through mechanisms that include CTLA-4 up-regulation and demonstrate a link between CD45 and CTLA-4 that depends on calcineurin-mediated signaling. They demonstrate also that CTLA-4 expression may be specifically targeted to enhance allograft acceptance.

Abatacept↗

PubMed2001

CD28-independent induction of experimental autoimmune encephalomyelitis.

Experimental autoimmune encephalomyelitis (EAE) is a T cell-mediated disease initiated by antigen-specific CD4(+) T cells. Signaling through CD28 is a critical second signal for activation of T cells, and CD28 knockout (CD28KO) mice have been reported to be resistant to induction of EAE. We now report that CD28KO mice have no intrinsic defect in mediating disease, because they developed EAE after passive transfer of primed T cells. After immunization, peripheral T cells from CD28KO mice were primed and developed memory phenotype, but had decreased antigen-specific IFN-gamma production as compared with cells from wild-type (WT) animals. Reimmunization of CD28KO mice brought out clinical disease and increased IFN-gamma production in vitro. Pathologically, there were cellular infiltrates in the central nervous system, in contrast to single-immunized mice. We show furthermore that blocking B7-1 or CTLA4, but not B7-2, in CD28KO mice induces disease after a single immunization. Thus, EAE can be induced in animals lacking CD28-dependent costimulation, suggesting that alternative costimulatory pathways were used. Blocking the OX40-OX40L costimulatory pathway differentially affected disease induction in CD28KO mice as compared with WT controls. Our data show that EAE may develop in the absence of CD28 T-cell costimulation. These findings have implications for therapies aimed at blocking costimulatory signals in autoimmune diseases.

Abatacept↗

PubMed2001

Regulatory functions of self-restricted MHC class II allopeptide-specific Th2 clones in vivo.

We studied T-cell clones generated from grafts of rejecting and tolerant animals and investigated the regulatory function of Th2 clones in vitro and in vivo. To prevent allograft rejection, we treated LEW strain recipient rats of WF strain kidney grafts with CTLA4Ig to block CD28-B7 costimulation. We then isolated epitope-specific T-cell clones from the engrafted tissue, using a donor-derived immunodominant class II MHC allopeptide presented by recipient antigen-presenting cells. Acutely rejected tissue from untreated animals yielded self-restricted, allopeptide-specific T-cell clones that produced IFN-gamma, whereas clones from tolerant animals produced IL-4 and IL-10. Adoptive transfer into naive recipients of Th1 clones, but not Th2 clones, induced alloantigen-specific delayed-type hypersensitivity (DTH) responses. In addition, Th2 clones suppressed DTH responses mediated by Th1 clones in vivo and blocked Th1 cell proliferation and IFN-gamma production in vitro. A pilot human study showed that HLA-DR allopeptide-specific T-cell clones generated from patients with chronic rejection secrete Th1 cytokines, whereas those from patients with stable graft function produce Th2 cytokines in response to donor-specific HLA-DR allopeptides. We suggest that self-restricted alloantigen-specific Th2 clones may regulate the alloimmune responses and promote long-term allograft survival and tolerance.

Animals↗

PubMed2001

B7-dependent T-cell costimulation in mice lacking CD28 and CTLA4.

To examine whether B7 costimulation can be mediated by a molecule on T cells that is neither CD28 nor CTLA4, we generated mice lacking both of these receptors. CD28/CTLA4(-/-) mice resemble CD28(-/-) mice in having decreased expression of T-cell activation markers in vivo and decreased T-cell proliferation in vitro, as compared with wild-type mice. Using multiple approaches, we find B7-dependent costimulation in CD28/CTLA4(-/-) mice. The proliferation of CD28/CTLA4(-/-) T cells is inhibited by CTLA4-Ig and by the use of antigen-presenting cells lacking both B7-1 and B7-2. CD28/CTLA4(-/-) T-cell proliferation is increased by exposure to Chinese hamster ovary cells transfected with B7-1 or B7-2. Finally, administration of CTLA4-Ig to CD28/CTLA4(-/-) cardiac allograft recipients significantly prolongs graft survival. These data support the existence of an additional receptor for B7 molecules that is neither CD28 nor CTLA4.

Abatacept↗

PubMed2001

Effect of targeted disruption of STAT4 and STAT6 on the induction of experimental autoimmune encephalomyelitis.

Experimental autoimmune encephalomyelitis (EAE) is mediated by myelin-specific CD4(+) T cells secreting Th1 cytokines, while recovery from disease is associated with expression of Th2 cytokines. Investigations into the role of individual cytokines in disease induction have yielded contradictory results. Here we used animals with targeted deletion of the STAT4 or STAT6 genes to determine the role of these signaling molecules in EAE. The STAT4 pathway controls the differentiation of cells into a Th1 phenotype, while the STAT6 pathway controls the differentiation of cells into a Th2 phenotype. We found that mice deficient in STAT4 are resistant to the induction of EAE, with minimal inflammatory infiltrates in the central nervous system. In contrast, STAT6-deficient mice, which have a predominantly Th1 phenotype, experience a more severe clinical course of EAE as compared with wild-type or STAT4 knockout mice. In addition, adoptive transfer studies confirm the regulatory functions of a Th2 environment in vivo. These novel data indicate that STAT4 and STAT6 genes play a critical role in regulating the autoimmune response in EAE.

Adoptive Transfer↗