CD28-mediated Costimulatory Signals Research Articles

Fish Uses CTLA-4 Immune Checkpoint to Suppress mTORC1-Controlled T-Cell Glycolysis and Immunity.

As an immune checkpoint, cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) suppresses the activation, proliferation, and effector function of T cells, thus preventing an overexuberant response and maintaining immune homeostasis. However, whether and how this immune checkpoint functions in early vertebrates remains unknown. In the current study, using a Nile tilapia (Oreochromis niloticus) model, we investigated the suppression of T cell response by CTLA-4 in bony fish. Tilapia CTLA-4 is constitutively expressed in lymphoid tissues, and its mRNA and protein expression in lymphocytes are upregulated following PHA stimulation or Edwardsiella piscicida infection. Blockade of CTLA-4 signaling enhanced T cell activation and proliferation but inhibited activation-induced T cell apoptosis, indicating that CTLA-4 negatively regulated T cell activation. In addition, blocking CTLA-4 signaling invivo increased the differentiation potential and cytotoxicity of T cells, resulting in an enhanced T cell response during E. piscicida infection. Tilapia CTLA-4 competitively bound the B7.2/CD86 molecule with CD28, thus antagonizing the CD28-mediated costimulatory signal of T cell activation. Furthermore, inhibition of mammalian/mechanistic target of rapamycin complex 1 (mTORC1) signaling, c-Myc, or glycolysis markedly impaired the CTLA-4 blockade-enhanced T cell response, suggesting that CTLA-4 suppressed the T cell response of tilapia by inhibiting mTORC1/c-Myc axis-controlled glycolysis. Overall, the findings indicate a detailed mechanism by which CTLA-4 suppresses T cell immunity in tilapia; therefore, we propose that early vertebrates have evolved sophisticated mechanisms coupling immune checkpoints and metabolic reprogramming to avoid an overexuberant T cell response.

Functional enhancement of mesothelin-targeted TRuC-T cells by a PD1-CD28 chimeric switch receptor

T cells expressing a mesothelin (MSLN)-specific T cell receptor fusion construct (TRuC®), called TC-210, have demonstrated robust antitumor activity in preclinical models of mesothelioma, ovarian cancer, and lung cancer. However, they are susceptible to suppression by the programmed cell death protein 1 (PD-1)/programmed cell death protein ligand 1 (PD-L1) axis and lack intrinsic costimulatory signaling elements. To enhance the function of anti-MSLN TRuC-T cells, chimeric switch receptors (CSRs) have been designed to co-opt the immunosuppressive PD-1/PD-L1 axis and to deliver a CD28-mediated costimulatory signal. Here, we report that coexpression of the PD1-CD28 CSR in TRuC-T cells enhanced T cell receptor signaling, increased proinflammatory effector cytokines, decreased anti-inflammatory cytokines, and sustained effector function in the presence of PD-L1 when compared with TC-210. Anti-MSLN TRuC-T cells engineered to coexpress PD1-CD28 CSRs comprising the ectodomain of PD-1 and the intracellular domain of CD28 linked by the transmembrane domain of PD-1 were selected for integration into an anti-MSLN TRuC-T cell therapy product called TC-510. In vitro, TC-510 showed significant improvements in persistence and resistance to exhaustion upon chronic stimulation by tumor cells expressing MSLN and PD-L1 when compared with TC-210. In vivo, TC-510 showed a superior ability to provide durable protection following tumor rechallenge, versus TC-210. These data demonstrate that integration of a PD1-CD28 CSR into TRuC-T cells improves effector function, resistance to exhaustion, and prolongs persistence. Based on these findings, TC-510 is currently being evaluated in patients with MSLN-expressing solid tumors.

Quantitative and Qualitative Deficits in Neonatal Lung-Migratory Dendritic Cells Impact the Generation of the CD8+ T Cell Response

CD103+ and CD11b+ populations of CD11c+MHCIIhi murine dendritic cells (DCs) have been shown to carry antigens from the lung through the afferent lymphatics to mediastinal lymph nodes (MLN). We compared the responses of these two DC populations in neonatal and adult mice following intranasal infection with respiratory syncytial virus. The response in neonates was dominated by functionally-limited CD103+ DCs, while CD11b+ DCs were diminished in both number and function compared to adults. Infecting mice at intervals through the first three weeks of life revealed an evolution in DC phenotype and function during early life. Using TCR transgenic T cells with two different specificities to measure the ability of CD103+ DC to induce epitope-specific CD8+ T cell responses, we found that neonatal CD103+ DCs stimulate proliferation in a pattern distinct from adult CD103+ DCs. Blocking CD28-mediated costimulatory signals during adult infection demonstrated that signals from this costimulatory pathway influence the hierarchy of the CD8+ T cell response to RSV, suggesting that limited costimulation provided by neonatal CD103+ DCs is one mechanism whereby neonates generate a distinct CD8+ T cell response from that of adults.

Antagonist properties of monoclonal antibodies targeting human CD28

Antagonist antibodies targeting CD28 have been proposed as an alternative to the use of CD80/86 antagonists to modulate T cell responses in autoimmunity and transplantation. Advantages would be the blockade of CD28-mediated co-stimulatory signals without impeding the co-inhibitory signals dependent on CD80 interactions with CTLA-4 and PD-L1 that are important for the control of immune responses and for the function of regulatory T cells. Anti-CD28 antibodies are candidate antagonists only if they prevent access to the CD80/86 ligands without simultaneously stimulating CD28 itself, a process that is believed to depend on receptor multimerization. In this study, we evaluated the impact of different formats of a potentially antagonist anti-human CD28 antibody on T cell activation. In particular, we examined the role of valency and of the presence of an Fc domain, two components that might affect receptor multimerization either directly or in the presence of accessory cells expressing Fc receptors. Among monovalent (Fab’, scFv), divalent (Fab’2), monovalent-Fc (Fv-Fc) and divalent-Fc (IgG) formats, only the monovalent formats showed consistent absence of induced CD28 multimerization and absence of associated activation of phosphoinositol-3-kinase, and clear antagonist properties in T cell stimulation assays. In contrast, divalent antibodies showed agonist properties that resulted in cell proliferation and cytokine release in an Fc-independent manner. Conjugation of monovalent antibodies with polyethylene glycol, α-1-antitrypsin or an Fc domain significantly extended their in vivo half-life without modifying their antagonist properties. In conclusion, these data indicate that monovalency is mandatory for maintaining the antagonistic activity of anti-CD28 monoclonal antibodies.

Antagonist properties of monoclonal antibodies to human CD28: role of valency and heavy-chain constant domain

Background Antagonist antibodies targeting CD28 have been proposed as an alternative to the use of CD80/86 antagonists to modulate T cell responses in autoimmunity and transplantation. Advantages would be the blockade of CD28mediated co-stimulatory signals without impeding co-inhibitory signals depending on CD80 interactions with CTLA-4 and PD-L1 that are important for the control of immune responses and for the function of regulatory T cells. Anti-CD28 antibodies are candidate antagonists only if they prevent access to the CD80/86 ligands without simultaneously stimulating CD28 itself, a process that is believed to depend on receptor multimerization.

THERAPEUTIC HOTLINE: Abatacept: our experience of use in two patients with refractory psoriasis and psoriatic arthritis

The B7 family of molecules on antigen presenting cells (APCs) regulate T cell activation. They deliver stimulatory signals through CD28 and inhibitory signals through CD152, or cytotoxic T lymphocyte-associated antigen-4 (CTLA-4). CTLA4Ig (abatacept) is a soluble chimeric protein consisting of the extracellular domain of human CD152 linked to the modified Fc portion of human IgG1. By binding to B7-1 (CD80) and B7-2 (CD86) molecules on APCs, CTLA4Ig blocks the CD28-mediated costimulatory signal for T cell activation. Success with abatacept has been noted in psoriasis. Abatacept was administered to two patients with refractory psoriasis and psoriatic arthritis after the patients had failed all conventional treatment methods. Both patients experienced very brief improvement in disease. The improvement, however, was not continued, and both patients were taken off the medication. The small patient population limits the extrapolation of the present authors' results to the larger population. Furthermore, the present authors' patients have very severe, refractory disease and do not adequately represent the majority of psoriasis patients. Although the present authors' patients demonstrated brief response to drug, this response was not sustained. No adverse events were reported in the present authors' patients.

Low-Dose Total Body Irradiation and Fludarabine Conditioning for HLA Class I-Mismatched Donor Stem Cell Transplantation and Immunologic Recovery in Patients with Hematologic Malignancies: A Multicenter Trial

HLA-mismatched grafts are a viable alternative source for patients without HLA-matched donors receiving ablative hematopoietic cell transplantation (HCT), although their use in reduced intensity conditioning (RIC) or nonmyeloablative (NMA) conditioning HCT has been not well established. Here, we extended HCT to recipients of HLA class I-mismatched grafts to investigate whether NMA conditioning can establish stable donor engraftment. Fifty-nine patients were conditioned with fludarabine (Flu) 90 mg/m(2) and 2 Gy total body irradiation (TBI), followed by immunosuppression with cyclosporine (CsA) 5.0 mg/kg twice a day and mycophenolate mofetil (MMF) 15 mg/kg 3 times a day for transplantation of granulocyte colony-stimulating factor (G-CSF)-mobilized peripheral blood stem cells (PBSCs) from related (n = 5) or unrelated donors (n = 54) with 1 antigen +/- 1 allele HLA class I mismatch or 2 HLA class I allele mismatches. Sustained donor engraftment was observed in 95% of the evaluable patients. The incidence of grade II-IV acute and extensive chronic graft-versus-host disease (aGVHD, cGVHD) was 69% and 41%, respectively. The cumulative probability of nonrelapse mortality (NRM) was 47% at 2 years. Two-year overall and progression-free survival (OS, PFS) was 29% and 28%, respectively. NMA conditioning with Flu and low-dose TBI, followed by HCT using HLA class I-mismatched donors leads to successful engraftment and long-term survival; however, the high incidence of aGVHD and NRM needs to be addressed by alternate GVHD prophylaxis regimens.

CD28-mediated costimulation impacts on the differentiation of DC vaccination-induced T cell responses

Costimulatory signals such as the ones elicited by CD28/B7 receptor ligation are essential for efficient T cell activation but their role in anti-tumour immune responses remains controversial. In the present study we compared the efficacy of DC vaccination-induced melanoma specific T cell responses to control the development of subcutaneous tumours and pulmonary metastases in CD28-deficient mice. Lack of CD28-mediated costimulatory signals accelerated tumour development in both model systems and also the load of pulmonary metastases was strongly increased by the end of the observation period. To scrutinize whether lack of CD28 signalling influences priming, homing or effector function of Trp-2(180-188)/K(b)-reactive T cells we investigated the characteristics of circulating and tumour infiltrating T cells. No difference in the frequency of Trp-2(180-188)/K(b)-reactive CD8+ T cells could be demonstrated among the cellular infiltrate of subcutaneous tumours after DC vaccination between both genotypes. However, the number of IFN-gamma-producing Trp-2-reactive cells was substantially lower in CD28-deficient mice and also their cytotoxicity was reduced. This suggests that CD28-mediated costimulatory signals are essential for differentiation of functional tumour-specific CD8+ T-effector cells despite having no impact on the homing of primed CD8+ T cells.

Differential Control of CD28-Regulated In Vivo Immunity by the E3 Ligase Cbl-b

The E3 ubiquitin ligase Casitas B cell lymphoma-b (Cbl-b) plays a critical role in the development of autoimmunity and sets the threshold for T cell activation. In the absence of Cbl-b, T cells stimulated via the TCR respond similarly to those that have received a CD28-mediated costimulatory signal, suggesting that the absence of Cbl-b substitutes for CD28-mediated costimulation. In this study, we show that loss of Cbl-b restores Ig class switching and germinal center formation in Vav1 mutant mice in response to an in vivo viral challenge. Genetic inactivation of Cbl-b also rescues impaired antiviral IgG production in CD28-mutant mice. Moreover, loss of CD28 results in disorganization of follicular dendritic cell clusters, which is also rescued by the Cbl-b mutation. Intriguingly, despite restored antiviral in vivo immunity and follicular dendritic cell clusters, loss of Cbl-b did not rescue germinal center formation in CD28-deficient mice. Mechanistically, in vivo vesicular stomatitis virus-induced IL-4 and IFN-gamma production and up-regulation of the inducible costimulatory molecule ICOS were dependent on CD28, and could not be rescued by the loss of Cbl-b. These data provide genetic evidence that CD28-dependent in vivo immune responses and Ig class switching can be genetically uncoupled from germinal center formation and ICOS induction by Cbl-b-Vav1-regulated signaling pathways.

Time course of gene expression after the injection of adenoviral vectors containing CTLA4IG gene

Introduction In vivo gene transfection using a recombinant adenoviral vector leads to diminished gene expression in a time-dependent manner that disappears within 4 weeks. CTLA4Ig blocks CD28-mediated costimulatory signal, and inhibits immune responses. We investigated the duration of transgene expression after administration of adenoviral vector containing CTLA4Ig gene (AdCTLA4Ig). Methods We injected 1 × 10 9 plaque forming units (pfu) of AdCTLA4Ig into rats (n = 7) via the tail vein. Thereafter, the blood samples were collected for assay of serum CTLA4Ig levels using enzyme-linked immunosorbent assay. Results The CTLA4Ig level reached the maximum (range, 65–86 μg/mL; average, 75 μg/mL) on days 3 to 5 after injection. Detectable levels of CTLA4Ig were observed up to 49 days. When we injected AdCTLA4Ig in combination with FTY720 administration, the maximum levels were higher and the detectable levels persisted longer. Conclusions Because directly injected adenoviral transgene expression had been reported to disappear between 21 to 30 days, we conclude that AdCTLA4Ig inhibits the immune response and prolongs the transgene (CTLA4Ig gene) expression. Some additional immunosuppressants, like FTY720, may be useful to enhance AdCTLA4Ig effects.

Activation requirements for the induction of CD4+CD25+ T cell suppressor function.

The in vivo differentiation/survival of CD4(+)CD25(+) T suppressor cells is dependent on IL-2 and CD28-mediated costimulatory signals. To determine the cytokine and costimulatory requirements for CD25(+) T cells in vitro, we established a two-stage culture system where CD25(+) T cells were activated in a primary culture. In the subsequent culture, activated CD4(+)CD25(+) cells were then mixed with responders in order to assess their suppressor function. Pre-culture of CD25(+) T cells with anti-CD3 alone resulted in poor survival and minimal induction of suppressor activity. Pre-culture in the presence of anti-CD3 and IL-2 or IL-4, but not IL-6, IL-7, IL-9, IL-10 or IL-15, resulted in proliferation of the CD25(+) cells and induction of potent suppressor function. Inhibition of the interaction of CD28 or cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) with CD80/CD86 in the pre-culture of CD4(+)CD25(+) cells did not prevent the induction of suppressor function. Furthermore, the inhibition of costimulatory signals did not inhibit the ability of fresh CD25(+) T cells to inhibit CD8(+) responders under conditions where activation of the responders was independent of CD80/CD86. These studies support the view that activation of CD25(+) T cells requires IL-2/IL-4 for their survival/differentiation into effector cells, but is independent of CD28/CTLA-4-mediated costimulation.

Tyrosine phosphorylation of a novel 100-kDa protein coupled to CD28 in resting human T cells is enhanced by a signal through TCR/CD3 complex.

For T cell activation, two signals are required, i.e., a T cell receptor (TCR)/CD3-mediated main signal and a CD28-mediated costimulatory signal. CD28 binds to its ligand (CD80 or CD86) and transduces the most important costimulatory signal. The cytoplasmic domain of the CD28 molecule, composed of 41 amino acids, does not contain any intrinsic enzyme activity. The cytoplasmic domain of CD28 is remarkably conserved among species and is associated with a number of signaling molecules that affect the main signal. We report here that a tyrosine phosphorylated 100-kDa protein (ppl00) was coupled to the CD28 cytoplasmic domain in Jurkat and human peripheral T cells. The pp100 was distinguished from other CD28 associated molecules such as Vav, STAT5, PI 3-kinase, Valosin-containing protein (VCP), Nucleolin, Gab2 (Grb2-associated binding protein 2), and STAT6. The tyrosine phosphorylation of pp100 coprecipitated with CD28 was enhanced by CD3 stimulation by the specific antibody, tyrosine phosphatase inhibitor and PKC activator. Tyrosine phosphorylation of pp100 was attenuated by the prior addition of PKC inhibitor. These findings indicate that pp100 is a novel tyrosine phosphorylated protein coupled to CD28 under continuous control of tyrosine phosphatases and might play a role in T cell activation augmented by a TCR/CD3-mediated main signal.

CD28 signaling augments Elk-1-dependent transcription at the c-fos gene during antigen stimulation.

Untransformed CD4(+) Th1 cells stimulated with Ag and APC demonstrated a dependence on B7- and CD28-mediated costimulatory signals for the expression and function of AP-1 proteins. The induction of transactivation by the c-fos gene regulator Elk-1 mirrored this requirement for TCR and CD28 signal integration. c-Jun N-terminal kinase (JNK) (but not extracellular signal-regulated kinase or p38) protein kinase activity was similarly inhibited by neutralizing anti-B7 mAbs. Blockade of JNK protein kinase activity with SB 202190 prevented both Elk-1 transactivation and c-Fos induction. These results identify a unique role for B7 costimulatory molecules and CD28 in the activation of JNK during Ag stimulation in Th1 cells, and suggest that JNK regulates Elk-1 transactivation at the c-fos gene to promote the formation of AP-1 complexes important to IL-2 gene expression.

Regulation of c-Jun NH(2)-terminal kinase (Jnk) gene expression during T cell activation.

The c-Jun NH(2)-terminal kinases (JNKs) are a group of mitogen-activated protein (MAP) kinases that participate in signal transduction events mediating specific cellular functions. Activation of JNK is regulated by phosphorylation in response to cellular stress and inflammatory cytokines. Here, we demonstrate that JNK is regulated by a second, novel mechanism. Induction of Jnk gene expression is required in specific tissues before activation of this signaling pathway. The in vivo and in vitro ligation of the T cell receptor (TCR) leads to induction of JNK gene and protein expression. TCR signals are sufficient to induce JNK expression, whereas JNK phosphorylation also requires CD28-mediated costimulatory signals. Therefore, both expression and activation contribute to the regulation of the JNK pathway to ensure proper control during the course of an immune response.

Prolonged cardiac allograft survival in rats systemically injected adenoviral vectors containing CTLA4Ig-gene.

CTLA4Ig, a soluble recombinant fusion protein that contains the extracellular domain of the CTLA4 and Fc portion of IgG1, strongly adheres to the B7 molecule to block CD28-mediated costimulatory signals and inhibits in vitro and in vivo immune responses. In vivo gene transfer using adenovirus vector achieves a high transfection rate into organ cells that usually contain adenoviral receptors. In this study, we investigated expression levels of the transfected gene and the survival times of the allografts in cardiac recipients systemically administered adenoviral vectors containing CTLA4Ig. Hearts from DA rats (RT-1a) were transplanted into a cervical location in LEW recipients (RT1(1)). The adenoviral vectors containing CTLA4Ig was injected via a recipient vein immediately after grafting. The serum level of CTLA4Ig reached to maximum at 51-93 microg/ml 3 to 7 days after gene-transfection and declined after 14 days, although detectable levels were observed up to 49 days. The median survival time of the allografts in the gene-transfected group were significantly prolonged (27 days) in compared to the control group (6 days). In addition, down-regulation of IL-2 and IFN-gamma mRNAs and persistence of IL-4 and IL-10 transcripts were observed in the graft infiltrating cells. The adenovirous-mediated CTLA4Ig gene transfer into a recipient liver by systemic administration resulted in remarkable prolongation of cardiac allograft survival. Its action mechanisms may be mediated by inhibition of CD28-associated signal transduction, reduction of Th1-type cytokine production, and continuous expression of Th2-type cytokines in the activating lymphocytes.

Effects of dietary docosahexaenoic acid on surface molecules involved in T cell proliferation

It is known that n-3 polyunsaturated fatty acids (PUFA) such as docosahexaenoic acid (DHA) suppress immunity as compared with n-6 PUFA such as linoleic acid (LA), but the mechanism involved in this phenomenon is still unclear. The present study was designed to assess the effect of dietary DHA on the surface molecules involved in T cell proliferation. Weanling male C57BL/6 mice were divided into four dietary groups that were fed a 10% fat diet for 4 weeks varying in amounts of DHA and LA. As the dietary DHA concentration increased, the surface expression of CD4 and CD8 on splenic T cells decreased, while that of CD28 increased. The surface expression of CD3, however, was invariable in all dietary groups. DNA synthesis of splenic T cells, induced by CD3 crosslinkage with anti-CD3ϵ monoclonal antibody in the presence of CD28-mediated costimulation, increased as the DHA concentration was elevated. These observations suggest that diets rich in DHA exert some of their immunomodulatory effects by a downregulation of surface expression of CD4 and CD8 and by an upregulation of CD28-mediated costimulatory signal.

Pyk2 is differentially regulated by beta1 integrin- and CD28-mediated co-stimulation in human CD4+ T lymphocytes.

Beta1 integrins can provide T cell co-stimulation, but little is known concerning their downstream signaling pathways. We found that Pyk2, a focal adhesion kinase-related tyrosine kinase, is regulated by beta1 integrin signaling in human T cells. Stimulation of Jurkat T cells with the alpha4beta1 integrin ligand VCAM-1 results in Pyk2 tyrosine phosphorylation, and combined stimulation with VCAM-1 and anti-CD3 mAb induces rapid and sustained synergistic Pyk2 phosphorylation. Studies with mAb suggest that in synergistic CD3- and alpha4beta1 integrin-mediated Pyk2 tyrosine phosphorylation, a major contribution of CD3-derived signals is independent of their effects on regulating integrin adhesion. Analysis of resting human CD4+ T cells confirmed the ability of CD3-derived signals to synergize with beta1 integrin-dependent signals in the induction of Pyk2 tyrosine phosphorylation. In addition, although CD28-mediated co-stimulatory signals were able to synergize with CD3-mediated signals in inducing ERK and JNK activation and secretion of IL-2 in the primary T cells, they did not contribute to the induction of Pyk2 phosphorylation. Taken together, these results indicate a potential role for Pyk2 in T cell co-stimulation mediated specifically by beta1 integrins.

Pyk2 is differentially regulated by β 1 integrin- and CD28-mediated co-stimulation in human CD4+ T lymphocytes

β1 integrins can provide T cell co-stimulation, but little is known concerning their downstream signaling pathways. We found that Pyk2, a focal adhesion kinase-related tyrosine kinase, is regulated by β1 integrin signaling in human T cells. Stimulation of Jurkat T cells with the α4β1 integrin ligand VCAM-1 results in Pyk2 tyrosine phosphorylation, and combined stimulation with VCAM-1 and anti-CD3 mAb induces rapid and sustained synergistic Pyk2 phosphorylation. Studies with mAb suggest that in synergistic CD3- and α4β1 integrin-mediated Pyk2 tyrosine phosphorylation, a major contribution of CD3-derived signals is independent of their effects on regulating integrin adhesion. Analysis of resting human CD4+ T cells confirmed the ability of CD3-derived signals to synergize with β1 integrin-dependent signals in the induction of Pyk2 tyrosine phosphorylation. In addition, although CD28-mediated co-stimulatory signals were able to synergize with CD3-mediated signals in inducing ERK and JNK activation and secretion of IL-2 in the primary T cells, they did not contribute to the induction of Pyk2 phosphorylation. Taken together, these results indicate a potential role for Pyk2 in T cell costimulation mediated specifically by β1 integrins.

Critical role of Fas/Fas ligand interaction in CD28-independent pathway of allogeneic murine hepatocyte rejection

Cytolytic induction of T cells requires both the T-cell receptor (TCR)-mediated antigenic stimulation and the CD28-mediated co-stimulatory signal. Blockade of the interactions between CD28 and its ligands, CD80 and CD86, prolongs the survival of allografts in some transplantation models. However, we found that allogeneic hepatocytes were completely rejected within 7 days after intrasplenic transplantation, even when treated with monoclonal antibodies (mAbs) against CD80 and CD86 (anti-CD80/86). Recent studies have shown that there are two main mechanisms of T-cell-mediated cytotoxicity, perforin-based and Fas-based ones. It has been shown that the liver is highly sensitive to induction of apoptosis by an agonistic anti-Fas mAb. We then investigated the role of the Fas/Fas ligand (FasL) system in the CD28-independent allogeneic hepatocyte rejection. With the anti-CD80/86 mAb treatment, hepatocytes from C57BL/6 lpr/lpr (B6 lpr) mice, which express little Fas antigen, could survive for 7 days after intrasplenic transplantation, and hepatocytes from C57BL/6 (B6) mice could also survive for 7 days in the spleen of C3H/ He gld/gld (C3H gld) mice, which express no functional FasL. CD28-independent induction of cytotoxicity against allogeneic hepatocytes was not observed when the effector cells were derived from C3H gld mice. These results indicated that the Fas/FasL system plays a critical role in the CD28-independent pathway of allogeneic hepatocyte rejection. (Hepatology 1997 Oct;26(4):944-8)

Critical role of Fas/Fas ligand interaction in CD28-independent pathway of allogeneic murine hepatocyte rejection

Cytolytic induction of T cells requires both the T-cell receptor (TCR)-mediated antigenic stimulation and the CD28-mediated co-stimulatory signal. Blockade of the interactions between CD28 and its ligands, CD80 and CD86, prolongs the survival of allografts in some transplantation models. However, we found that allogeneic hepatocytes were completely rejected within 7 days after intrasplenic transplantation, even when treated with monoclonal antibodies (mAbs) against CD80 and CD86 (anti-CD80/86). Recent studies have shown that there are two main mechanisms of T-cell-mediated cytotoxicity, perforin-based and Fas-based ones. It has been shown that the liver is highly sensitive to induction of apoptosis by an agonistic anti-Fas mAb. We then investigated the role of the Fas/Fas ligand (FasL) system in the CD28-independent allogeneic hepatocyte rejection. With the anti-CD80/86 mAb treatment, hepatocytes from C57BL/6 lpr/lpr (B6 lpr) mice, which express little Fas antigen, could survive for 7 days after intrasplenic transplantation, and hepatocytes from C57BL/6 (B6) mice could also survive for 7 days in the spleen of C3H/ He gld/gld (C3H gld) mice, which express no functional FasL. CD28-independent induction of cytotoxicity against allogeneic hepatocytes was not observed when the effector cells were derived from C3H gld mice. These results indicated that the Fas/FasL system plays a critical role in the CD28-independent pathway of allogeneic hepatocyte rejection.