Donor-Reactive CD8 Memory T Cells Infiltrate Cardiac Allografts Within 24-h Posttransplant in Naive Recipients

Normal immune responses stimulated by pathogenic and environmental antigens generate memory T cells that react with donor antigens and no currently used immunosuppressive drug completely inhibits memory T-cell function. While donor-reactive memory T cells clearly compromise graft outcomes, mechanisms utilized by memory T cells to promote rejection are largely unknown. In this study, we investigated how early endogenous memory cells infiltrate and express effector function in cardiac allografts. Endogenous CD8 memory T cells in nonsensitized recipients distinguish syngeneic versus allogeneic cardiac allografts within 24 h of reperfusion. CD8-dependent production of IFN-γ and CXCL9/Mig was observed 24 to 72 h posttransplant in allografts but not isografts. CXCL9 was produced by donor cells in response to IFN-γ made by recipient CD8 T cells reactive to donor class I major histocompatibility complex (MHC) molecules. Activated CD8 T cells were detected in allografts at least 3 days before donor-specific effector T cells producing IFN-γ were detected in the recipient spleen. Early inflammation mediated by donor-reactive CD8 memory T cells greatly enhanced primed effector T-cell infiltration into allografts. These results suggest that strategies for optimal inhibition of alloimmunity should include neutralization of infiltrating CD8 memory T cells within a very narrow window after transplantation.     

IFN-γ Decreases CTL Generation by Limiting IL-2 Production: A Feedback Loop Controlling Effector Cell Production

IFN-γ is produced by cytotoxic T lymphocytes (CTL) but can also decrease CTL generation. We used IFN-γ-R1-deficient (GRKO) and IFN-γ-deficient (GKO) mice to study the effects of IFN-γ in MLC on the generation of CTL activity and CTL number, IL-2 production and cell proliferation. CTL activity was increased in MLC when GRKO responders or GKO stimulators and responders were used, compared to wild-type (WT) MLC. The number of cells displaying the CTL phenotype (CD3⁺, CD8⁺, CD25⁺) was also increased, accompanied by increased IL-2 production and proliferation. Combinations of WT or GRKO CD4+ T cells with WT or GRKO CD8+ T cells as responders showed that IFN-γ mostly affects CD4+ T cells to limit CTL generation. Intracellular staining indicated that IL-2 production was largely by CD4+ T cells. Moreover, addition of IL-2 to WT responders mimicked GKO CTL generation and activity, whereas neutralizing IL-2 decreased CTL activity in GRKO and WT responders. Thus IFN-γ reduces CTL generation in alloimmune responses largely by limiting proliferation of IL-2 producing CD4+ T cells. This creates a feedback loop in which effectors produce IFN-γ that limits IL-2 production which in turn limits CTL generation.     

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.     

Prolonged survival in rat liver transplantation with mouse monoclonal antibody against an inducible costimulator (ICOS)

BACKGROUND: An inducible costimulator (ICOS), a recently identified costimulatory receptor with a close structural homology to CD28 and CTLA4, is expressed on activated T cells. Interaction with its ligand on antigen-presenting cells stimulates T-cell proliferation to produce a different spectrum of cytokine. The inhibition of ICOS-mediated signal transduction by an anti-ICOS antibody is considered to be capable of protecting against graft rejection in organ transplantation. METHODS: An anti-rat ICOS antibody was intravenously administered into recipients of dark Agouti-to-Lewis liver transplantations. The recipient lymphocytes from mesenteric lymph nodes were harvested on day 7 after transplantation for fluorescence-activated cell sorting analysis, and tissue specimens from the grafts were removed for histologic evaluation. Antigen-specific T-cell proliferation responses were assessed in vitro with anti-ICOS antibody. RESULTS: Monotherapy with the antibody significantly prolonged the graft survival time by inhibiting T-cell activation and its proliferation response. The graft-infiltrating cells, both CD4 and CD8 T cells, were not completely reduced even when rats were administered the antibody, whereas the expression of ICOS almost completely disappeared in these cells. CONCLUSIONS: T-cell activation through the ICOS costimulatory pathway plays an important role in graft rejection, and manipulating its pathway is an effective method for modulating transplantation immunity.     

ICOS-Dependent and -Independent Functions of Memory CD4 T Cells in Allograft Rejection

Donor-reactive memory T cells undermine the survival of transplanted organs through multiple pathways. We have previously reported that memory CD4 T cells resist treatment with anti-CD154 antibody and donor-specific transfusion (DST/MR1) and promote cardiac allograft rejection via generation of effector CD4 T cells and alloantibody. We hypothesized that the helper functions of memory CD4 T cells are independent of T-cell costimulation through CD154 but instead are regulated by alternative costimulatory pathways. This study investigated how blocking ICOS/B7RP-1 interactions affects functions of donor-reactive memory CD4 T cells. Treatment with blocking anti-ICOS mAb synergized with DST/MR1 and prolonged mouse cardiac allograft survival despite the presence of donor-reactive memory CD4 T cells. While blocking ICOS did not diminish the expansion of preexisting memory CD4 T cells or the induction of allospecific effector T cells, it did inhibit recruitment of the activated memory and effector T cells into the graft. In addition, anti-ICOS mAb treatment in combination with DST/MR1 prevented help provided by memory CD4 T cells for production of donor-specific IgG antibody. These results demonstrate the potential efficacy of ICOS blockade in sensitized transplant patients and provide the foundation for rational use of ICOS blockade in combination with other graft-prolonging strategies.     

他克莫司对肝移植受者外周血T淋巴细胞亚群及共刺激分子表达的影响

目的 探讨他克莫司（FK506）对肝移植受者外周血T淋巴细胞亚群及其表面共刺激分子表达的影响。方法 采用荧光标记单克隆抗体和流式细胞技术，测定术后采用FK506治疗的肝移植受者（FK506治疗组）在用FK506后1、2、3、4周时的外周血T淋巴细胞亚群及其表面共刺激分子CD28、CD152和ICOS分子的表达情况，以健康志愿者（健康对照组）和患终末期肝脏疾病拟行肝移植者（肝病对照组）为对照。结果 CD3^＋T淋巴细胞在各组间的差异均无统计学意义（P〉0．05）。经FK506治疗后，肝移植患者的CD4^＋T淋巴细胞逐渐减少，CD8^＋T淋巴细胞逐渐增加，并恢复至健康对照组水平（P〉0．05）。FK506治疗组T淋巴细胞亚群表面CD28分子和ICOS分子表达逐渐下降，并明显低于健康对照组（P〈0．05），而CD152分子表达增加，且明显高于健康对照组（P〈0．05）；其ICOS分子表达水平的下降晚于CD28分子，CD4^＋CD28^＋T淋巴细胞、CD8^＋CD28^＋T淋巴细胞和CD4^＋ICOS^＋T淋巴细胞均呈现相近的变化规律。结论 FK506能迅速纠正移植受者T淋巴细胞亚群紊乱，并抑制正性共刺激分子CD28和ICOS的表达，促进负性共刺激分子CD152的表达。     

Contribution of Naïve and Memory T-Cell Populations to the Human Alloimmune Response

T-cell alloimmunity plays a dominant role in allograft rejection. The precise contribution of naïve and memory T cells to this response however remains unclear. To address this question, we established an ex vivo flow-cytometric assay that simultaneously measures proliferation, precursor frequency and effector molecule (IFNγ, granzyme B/perforin) production of alloreactive T cells. By applying this assay to peripheral blood mononuclear cells from healthy volunteers, we demonstrate that the CD4⁺ and CD8⁺ populations mount similar proliferative responses and contain comparable frequencies of alloreactive precursors. Effector molecule expression, however, was significantly higher among CD8⁺ T cells. Analysis of sorted naïve and memory T cells showed that alloreactive precursors were equally present in both populations. The CD8⁺ effector and terminally differentiated effector memory subsets contained the highest proportion of granzyme B/perforin after allostimulation, suggesting that these cells present a significant threat to transplanted organs. Finally, we demonstrate that virus-specific lymphocytes contribute significantly to the alloresponse in certain responder–stimulator HLA combinations, underscoring the importance of T-cell cross-reactivity in alloimmunity. These results provide a quantitative assessment of the roles of naïve and memory T-cell subsets in the normal human alloimmune response and establish a platform for measuring T-cell alloreactivity pre- and posttransplantation.     

T cells in Cardiac Allograft Vasculopathy Are Skewed to Memory Th-1 Cells in the Presence of a Distinct Th-2 Population

Cardiac allograft vasculopathy (CAV) in heart transplantation (HTx) patients remains the major complication for long-term survival, due to concentric neointima hyperplasia induced by infiltrating mononuclear cells (MNC). Previously, we showed that activated memory T-helper-1 (Th-1) cells are the major component of infiltrating MNC in coronary arteries with CAV. In this study, a more detailed characterization of the MNC in human coronary arteries with CAV (n = 5) was performed and compared to coronary arteries without CAV (n = 5), by investigating MNC markers (CD1a, DRC-1, CD3, CD20, CD27, CD28, CD56, CD68, CD69, FOXP3 and HLA-DR), cytokines (IL-1A, 2, 4, 10, 12B, IFN-γ, and TGF-β1), and chemokine receptors (CCR3, CCR4, CCR5, CCR7, CCR8, CXCR3 and CX3CR1) by immunohistochemical double-labeling and quantitative PCR on mRNA isolated from laser microdissected layers of coronary arteries. T cells in the neointima and adventitia of CAV were skewed toward an activated memory Th-1 phenotype, but in the presence of a distinct Th-2 population. FOXP3 positive T cells were not detected and production of most cytokines was low or absent, except for IFN-γ, and TGF-β. This typical composition of T-helper cells and especially production of IFN-γ and TGF-β may play an important role in the proliferative CAV reaction.     

Quantification of immunosuppression by flow cytometric measurement of intracellular cytokine synthesis

Abstract The availibility of a method to measure the effects of drugs on immune reactivity should be helpful in optimizing treatment after organ transplantation. Since cyclosporine A (CSA) interferes with activation of T cells and cytokine synthesis, production of IL-2 and IFN-γ might constitute a marker of this drug's effects. We measured the capacity for mitogen-stimulated production of these cytokines in whole blood by using immunostaining of intracellular and membrane antigens, followed by flow cytometry. The percentage of CD4⁺ T cells producing IL-2 or IFN-γ was strongly reduced in 20 transplant patients compared with 24 healthy controls. The capacity for IL-2 production of CD4⁺ and CD8⁺ cells correlated inversely with CSA blood levels ( P values 0.0087 and 0.0396, respectively). IFN-γ production by CD4⁺ T cells showed a negative correlation with the prednisolone dose ( P = 0.0175) and, for the CD8⁺ subset, with CSA trough levels ( P = 0.0023). These data show that inhibition of T cell cytokine synthesis by CSA and prednisolone can be quantified.     

Morbidly obese human subjects have increased peripheral blood CD4+ T cells with skewing toward a Treg- and Th2-dominated phenotype

Obesity is associated with local T-cell abnormalities in adipose tissue. Systemic obesity-related abnormalities in the peripheral blood T-cell compartment are not well defined. In this study, we investigated the peripheral blood T-cell compartment of morbidly obese and lean subjects. We determined all major T-cell subpopulations via six-color flow cytometry, including CD8+ and CD4+ T cells, CD4+ T-helper (Th) subpopulations, and natural CD4+CD25+FoxP3+ T-regulatory (Treg) cells. Moreover, molecular analyses to assess thymic output, T-cell proliferation (T-cell receptor excision circle analysis), and T-cell receptor-β (TCRB) repertoire (GeneScan analysis) were performed. In addition, we determined plasma levels of proinflammatory cytokines and cytokines associated with Th subpopulations and T-cell proliferation. Morbidly obese subjects had a selective increase in peripheral blood CD4+ naive, memory, natural CD4+CD25+FoxP3+ Treg, and Th2 T cells, whereas CD8+ T cells were normal. CD4+ and CD8+ T-cell proliferation was increased, whereas the TCRB repertoire was not significantly altered. Plasma levels of cytokines CCL5 and IL-7 were elevated. CD4+ T-cell numbers correlated positively with fasting insulin levels. The peripheral blood T-cell compartment of morbidly obese subjects is characterized by increased homeostatic T-cell proliferation to which cytokines IL-7 and CCL5, among others, might contribute. This is associated with increased CD4+ T cells, with skewing toward a Treg- and Th2-dominated phenotype, suggesting a more anti-inflammatory set point.     

Acute lung transplant rejection is associated with localized increase in T-cell IFNgamma and TNFalpha proinflammatory cytokines in the airways

BACKGROUND: Allograft rejection remains a major cause of morbidity and mortality after lung transplantation and is associated with increased gene expression for proinflammatory cytokines. T cells are a major cell type involved in graft rejection. There have been no previous studies of cytokine production by T cells from blood, bronchoalveolar lavage (BAL), and intraepithelial T cells from bronchial brushings (BB) during rejection episodes; we hypothesized that T-cell proinflammatory cytokines would be increased in the airways during rejection episodes despite standard immunosuppression regimens. METHOD: To investigate changes in cytokine profiles during rejection episodes, whole blood, BAL, and BB from stable lung transplant patients and those with acute rejection were stimulated in vitro and intracellular cytokine production by CD8- (CD4+) and CD8+ T-cell subsets determined using multiparameter flow cytometry. RESULTS: Transforming growth factor (TGF)-beta was significantly decreased in blood CD4+ and CD8+ T cells while interferon (IFN)-gamma and tumor necrosis factor (TNF)-alpha were significantly increased in BAL CD4+ and CD8+ T cells in patients with evidence of rejection. There was no change in CD4:CD8, interleukin (IL)-2, or IL-4 between stable and rejecting groups. CONCLUSIONS: Acute lung transplant rejection is associated with decreased intracellular T-cell TGFbeta in blood and increased intracellular IFNgamma and TNFalpha in BAL CD4+ and CD8+ T cells. Drugs that effectively reduce airway T-cell IFNgamma and TNFalpha proinflammatory cytokine production may improve current protocols for reducing acute graft rejection in lung transplant patients.     

Primed CD8+ T-Cell Responses to Allogeneic Endothelial Cells Are Controlled by Local Complement Activation

CD8 T cells primed by transplantation recognize allogeneic class I MHC molecules expressed on graft vascular endothelium and contribute to allograft injury. We previously showed that immune cell-derived complement activation fragments are integral to T cell activation/expansion. Herein we tested the impact of local complement production/activation on T cell/endothelial cell (EC) interactions. We found that proinflammatory cytokines upregulated alternative pathway complement production by ECs, yielding C5a. We further found that ECs deficient in the cell surface C3/C5 convertase regulator decay accelerating factor (DAF, CD55) induced greater CD8 T-cell proliferation and more IFNγ⁺ and perforin⁺ effector cells than wild-type (WT) ECs. Allogeneic C3^−/− EC induced little or no CD8 responses. Abrogation of responses following C5a receptor (C5aR) blockade, or augmentation following addition of recombinant C5a demonstrated that the effects were mediated through T-cell-expressed-C5aR interactions. Analyses of in vivo CD8 cell responses to transplanted heart grafts deficient in EC DAF showed similar augmentation. The findings reveal that EC-derived complement triggers secondary CD8 T-cell differentiation and expansion and argue that targeting complement and/or C5aR could limit T-cell-mediated graft injury.     

LFA‐1 Antagonism Inhibits Early Infiltration of Endogenous Memory CD8 T Cells into Cardiac Allografts and Donor‐Reactive T Cell Priming

Alloreactive memory T cells are present in virtually all transplant recipients due to prior sensitization or heterologous immunity and mediate injury undermining graft outcome. In mouse models, endogenous memory CD8 T cells infiltrate MHC‐mismatched cardiac allografts and produce IFN‐γ in response to donor class I MHC within 24 h posttransplant. The current studies analyzed the efficacy of anti‐LFA‐1 mAb to inhibit early CD8 T cell cardiac allograft infiltration and activation. Anti‐LFA‐1 mAb given to C57BL/6 6 (H‐2^b) recipients of A/J (H‐2^a) heart grafts on days –1 and 0 completely inhibited CD8 T cell allograft infiltration, markedly decreased neutrophil infiltration and significantly reduced intragraft expression levels of IFN‐γ‐induced genes. Donor‐specific T cells producing IFN‐γ were at low/undetectable numbers in spleens of anti‐LFA‐1 mAb treated recipients until day 21. These effects combined to promote substantial prolongation (from day 8 to 27) in allograft survival. Delaying anti‐LFA‐1 mAb treatment until days 3 and 4 posttransplant did not inhibit early memory CD8 T cell infiltration and proliferation within the allograft. These data indicate that peritransplant anti‐LFA‐1 mAb inhibits early donor‐reactive memory CD8 T cell allograft infiltration and inflammation suggesting an effective strategy to attenuate the negative effects of heterologous immunity in transplant recipients.     

Memory T-Cell Predominance Following T-Cell Depletional Therapy Derives from Homeostatic Expansion of Naive T Cells

T-cell depletion reportedly leads to alterations in the T-cell compartment with predominant survival of memory phenotype CD4 T cells. Here, we asked whether the prevalence of memory T cells postdepletion results from their inherent resistance to depletion and/or to the homeostatic expansion of naive T cells and their phenotypic conversion to memory, which is known to occur in lymphopenic conditions. Using a 'mosaic memory' mouse model with trackable populations of alloreactive memory T cells, we found that treatment with murine antithymocyte globulin (mATG) or antilymphocyte serum (ALS) effectively depleted alloreactive memory CD4 T cells, followed by rapid homeostatic proliferation of endogenous CD4 T cells peaking at 4 days postdepletion, with no homeostatic advantage to the antigen-specific memory population. Interestingly, naive (CD44lo) CD4 T cells exhibited the greatest increase in homeostatic proliferation following mATG treatment, divided more extensively compared to memory (CD44hi) CD4 T cells and converted to a memory phenotype. Our results provide novel evidence that memory CD4 T cells are susceptible to lymphodepletion and that the postdepletional T-cell compartment is repopulated to a significant extent by homeostatically expanded naive T cells in a mouse model, with important important implications for immune alterations triggered by induction therapy.     

Acceptance of islet allografts in the liver of mice by blockade of an inducible costimulator

BACKGROUND: An inducible costimulator (ICOS) has been found to be a novel costimulator for T-cell activation, although its precise role in transplant immunobiology remains unclear. This study determined whether ICOS plays an essential role in rejection of intrahepatic islet allografts in streptozotocin-induced diabetic mice. METHODS: Mononuclear cells in the liver of mice were isolated and examined by flow cytometry with respect to expression of ICOS in association with rejection, and the effects of in vivo treatment with an anti-ICOS antibody on survival of intrahepatic islet allografts were determined.RESULTSFlow cytometric analysis of mononuclear cells in the liver of normal untreated mice revealed that ICOS is expressed on CD4+CD3int natural killer T cells. The expression of ICOS was up-regulated on CD4+CD3bright T cells and expanded CD8 T cells in the liver in association with rejection. Posttransplant short-term administration of anti-ICOS antibody alone produced a significant prolongation of islet allograft survival. Administration of the antibody in conjunction with a subtherapeutic regimen of FK506 prevented rejection, leading to the acceptance of islet allografts. CONCLUSION: ICOS has an essential role in rejection of intrahepatic islet allografts and the blockade of ICOS interaction might be a novel approach for preventing islet allograft rejection.     

可诱导共刺激分子与人IgG Fc融合蛋白诱导同种免疫低应答的体内外实验

目的 探讨真核表达人可诱导共刺激分子(ICOS)与人IgG Fc融合蛋白(ICOS-Ig融合蛋白)在体内外对同种免疫应答的影响.方法 构建ICOS-Ig融合蛋白表达载体,在CHO细胞中表达并纯化ICXIS-Ig融合蛋白.以Balb/c小鼠脾细胞为反应细胞,经Co60照射灭活的(257BL/6小鼠脾细胞为刺激细胞,进行初次混合淋巴细胞反应(MLR),MLR体系中分别加入50μ,/ml IOOS-Ig融合蛋白(ICOS-Ig组)或对照IgG(IgG组),采用氚标记胸腺嘧啶脱氧核苷(3H-TdR)掺入法检测反应细胞的增殖情况,酶联免疫吸附试验(ELSA)检测培养上清液中自细胞介素(IL)2、4、10以及γ干扰素(IFN-γ)的含量.收集初次MLR细胞,与灭活的C57BL/6小鼠脾细胞或C3H小鼠脾细胞共培养,进行再次MLR,检测指标同初次MLR.以Co60照射Balb/c小鼠,经尾静脉输注用羟基荧光素二醋酸盐琥珀酰亚胺脂(CFSE)标记的C57BL/6小鼠脾细胞,每天腹腔注射ICOS-Ig融合蛋白0.2 mg(IOOS-Ig组)、IgG(对照IgG组)或环孢素A(CsA组),3 d后取Balb/c小鼠脾细胞,流式细胞仪测定CFSE荧光强度以判断同种T淋巴细胞的体内增殖情况.结果 初次MLR显示,ICOS-Ig组同种T淋巴细胞活化增殖的抑制率为(58±8)%,其培养上清液中IFN-γ的水平明显高于IgG组(P<0.05).再次MLR显示,IOOS-Ig融合蛋白能特异性抑制C57BL/6小鼠脾细胞所致的细胞增殖,抑制率为(42±8)%,IL-4和Ⅱ-10的分泌受到抑制,而IFN-γ7的分泌增加;ICOS-Ig融合蛋白并不抑制第三方细胞所致的细胞增殖.体内实验显示,ICOS-Ig组和CsA组的CFSE荧光强度明显强于空白对照组和对照IgG组(P<0.05),而联合处理组CFSE荧光强度强于ICOS-Ig组和CsA组(P<0.05).结论 ICOS-Ig融合蛋白在体内外均可抑制同种T淋巴细胞的活化增殖,且这种作用具有特异性.     

Peritransplant VLA‐4 blockade inhibits endogenous memory CD8 T cell infiltration into high‐risk cardiac allografts and CTLA‐4Ig resistant rejection

Recipient endogenous memory CD8 T cells expressing reactivity to donor class I MHC infiltrate MHC‐mismatched cardiac allografts within 24 hours after reperfusion and express effector functions mediating graft injury. The current study tested the efficacy of Very Late Antigen‐4 (VLA‐4) blockade to inhibit endogenous memory CD8 T cell infiltration into cardiac allografts and attenuate early posttransplant inflammation. Peritransplant anti‐VLA‐4 mAb given to C57BL6 (H‐2^b) recipients of AJ (H‐2^a) heart allografts completely inhibited endogenous memory CD4 and CD8 T cell infiltration with significant decrease in macrophage, but not neutrophil, infiltration into allografts subjected to either minimal or prolonged cold ischemic storage (CIS) prior to transplant, reduced intra‐allograft IFN‐γ‐induced gene expression and prolonged survival of allografts subjected to prolonged CIS in CTLA‐4Ig treated recipients. Anti‐VLA‐4 mAb also inhibited priming of donor‐specific T cells producing IFN‐γ until at least day 7 posttransplant. Peritransplant anti‐VLA plus anti‐CD154 mAb treatment similarly prolonged survival of allografts subjected to minimal or increased CIS prior to transplant. Overall, these data indicate that peritransplant anti‐VLA‐4 mAb inhibits early infiltration memory CD8 T cell infiltration into allografts with a marked reduction in early graft inflammation suggesting an effective strategy to attenuate negative effects of heterologous alloimmunity in recipients of higher risk grafts.     

Inducible Co-Stimulator (ICOS) in transplantation: A review

Prevention of T cell activation is one of the goals of successful organ and tissue transplantation. Blockade of T cell co-stimulation, particularly of the CD28:B7 interaction, has been shown to prolong graft survival. Inducible Co-Stimulator (ICOS) is the third member of the B7 family and here we review the literature on ICOS, its receptor (B7RP-1), and blockade of this pathway in transplant models. ICOS:B7RP-1 are a single receptor:ligand pair with a loss of function of either being implicated in some autoimmune diseases. ICOS has multiple functions, related to its constitutive expression on B cells and activated T cells. In in vitro transplant models, ICOS:B7RP-1 blockade has produced mixed results as to its ability to modulate lymphocyte proliferation. Several in vivo transplant models demonstrate varying degrees of success in prolonging graft survival. Timing and dose of treatment appear important, and combination with other immunosuppressive treatments may also be of benefit. As ICOS has multiple functions, it may be that the observed variable results are due to inadvertent inactivation of graft protective functions. If these barriers can be overcome, ICOS:B7RP-1 blockade could provide an important target for future immunosuppression regimens.     

Striking Dichotomy of PD-L1 and PD-L2 Pathways in Regulating Alloreactive CD4+ and CD8+ T Cells In Vivo

Programmed death-1 (PD-1) is a recently identified coinhibitory molecule that belongs to the CD28 superfamily. PD-1 has two ligands PD-L1 and PD-L2. There is some evidence that PD-L1 and PD-L2 serve distinct functions, but their exact function in alloimmunity remains unclear. In the present study, we used a GVHD-like model that allows detailed analyses of T-cell activation at a single cell level in vivo to examine the role of PD-1/PD-L1 and PD-1/PD-L2 interactions in regulating proliferation of CD4(+) and CD8(+) T cells in response to alloantigen stimulation. We found that both CD4(+) and CD8(+) T cells proliferated vigorously in vivo and that PD-L1 and PD-L2 exhibit strikingly different effect on T-cell proliferation. While blocking PD-L1 did not affect the in vivo proliferation of CD4(+) and CD8(+) T cells regardless of CD28 costimulation, blocking PD-L2 resulted in a marked increase in the responder frequency of CD8(+) T-cells in vivo. The effect of PD-L2 on the CD8(+) T-cell proliferation is regulated by CD28 costimulation and by the CD4(+) T cells. We conclude that PD-L1 and PD-L2 function differently in regulating alloreactive T-cell activation in vivo, and PD-L2 is predominant in this model in limiting alloreactive CD8(+) T-cell proliferation.