The P2X7 receptor antagonist Brilliant Blue G reduces serum human interferon-γ in a humanized mouse model of graft-versus-host disease.

Graft-versus-host disease (GVHD) remains a major problem after allogeneic haematopoietic stem cell transplantation, a curative therapy for haematological malignancies. Previous studies have demonstrated a role for the adenosine triphosphate (ATP)-gated P2X7 receptor channel in allogeneic mouse models of GVHD. In this study, injection of human peripheral blood mononuclear cells (PBMCs) into immunodeficient non-obese diabetic-severe combined immunodeficiency-interleukin (NOD-SCID-IL)-2Rγnull (NSG) mice established a humanized mouse model of GVHD. This model was used to study the effect of P2X7 blockade in this disease. From five weeks post-PBMC injection, humanized mice exhibited clinical signs and histopathology characteristic of GVHD. The P2X7 antagonist, Brilliant Blue G (BBG), blocked ATP-induced cation uptake into both murine and human cells in vitro. Injection of BBG (50 mg/kg) into NSG mice did not affect engraftment of human leucocytes (predominantly T cells), or the clinical score and survival of mice. In contrast, BBG injection reduced circulating human interferon (IFN)-γ significantly, which was produced by human CD4+ and CD8+ T cells. BBG also reduced human T cell infiltration and apoptosis in target organs of GVHD. In conclusion, the P2X7 antagonist BBG reduced circulating IFN-γ in a humanized mouse model of GVHD supporting a potential role for P2X7 to alter the pathology of this disease in humans.

Exacerbation of spontaneous autoimmune nephritis following regulatory T cell depletion in B cell lymphoma 2-interacting mediator knock-out mice.

Regulatory T cells (Tregs ) have been recognized as central mediators for maintaining peripheral tolerance and limiting autoimmune diseases. The loss of Tregs or their function has been associated with exacerbation of autoimmune disease. However, the temporary loss of Tregs in the chronic spontaneous disease model has not been investigated. In this study, we evaluated the role of Tregs in a novel chronic spontaneous glomerulonephritis model of B cell lymphoma 2-interacting mediator (Bim) knock-out mice by transient depleting Tregs . Bim is a pro-apoptotic member of the B cell lymphoma 2 (Bcl-2) family. Bim knock-out (Bim-/- ) mice fail to delete autoreactive T cells in thymus, leading to chronic spontaneous autoimmune kidney disease. We found that Treg depletion in Bim-/- mice exacerbated the kidney injury with increased proteinuria, impaired kidney function, weight loss and greater histological injury compared with wild-type mice. There was a significant increase in interstitial infiltrate of inflammatory cells, antibody deposition and tubular damage. Furthermore, the serum levels of cytokines interleukin (IL)-2, IL-4, IL-6, IL-10, IL-17α, interferon (IFN)-γ and tumour necrosis factor (TNF)-α were increased significantly after Treg depletion in Bim-/- mice. This study demonstrates that transient depletion of Tregs leads to enhanced self-reactive T effector cell function followed by exacerbation of kidney disease in the chronic spontaneous kidney disease model of Bim-deficient mice.

Optimizing Outcomes in Pediatric Renal Transplantation Through the Australian Paired Kidney Exchange Program.

Kidney paired donation (KPD) programs offer the opportunity to enable living kidney donation when immunological and other barriers prevent safe directed donation. Children are likely to require multiple transplants during their lifetime; therefore, high-level histocompatibility and organ quality matching are key priorities. Details are given for a cohort of seven pediatric renal transplantations performed through the Australian Kidney Exchange (AKX), including barriers to alternative transplantation and outcomes after KPD. Reasons for entering the KPD program were preformed donor-specific antibodies to their registered donor in five cases, ABO mismatch, and avoidance of the risk of exposure to hepatitis B virus. Four recipients were highly sensitized. All patients received transplants with organs of lower immunological risk compared with their registered donors. HLA eplet mismatch scores were calculated for donor-recipient pairs; three patients had improved eplet mismatch load with AKX donor compared with their registered donor. All grafts are functioning, with a mean estimated glomerular filtration rate of 77 mL/min/1.73 m2 (range 46-94 mL) and a follow-up range of 8-54 months, and no patient experienced clinical or histological rejection. KPD is a viable strategy to overcome many barriers to living donation for pediatric patients who have an otherwise suitable donor and provides an opportunity to minimize immunological risks.

IL-17 deficiency attenuates allograft injury and prolongs survival in a murine model of fully MHC-mismatched renal allograft transplantation.

IL-17 is a pro-inflammatory cytokine implicated in the pathogenesis of inflammatory and autoimmune diseases. However the role of IL-17 in renal allograft rejection has not been fully explored. Here, we investigate the impact of IL-17 in a fully MHC-mismatched, life-sustaining, murine model of kidney allograft rejection using IL-17 deficient donors and recipients (IL-17(-/-) allografts). IL-17(-/-) allografts exhibited prolonged survival which was associated with reduced expression of the Th1 cytokine IFN-γ and histological attenuation of acute and chronic allograft rejection, as compared to wild-type allograft recipients. Results were confirmed in WT allograft recipients treated with an IL-17 blocking antibody. Subsequent experiments using either donors or recipients deficient in IL-17 showed a trend towards prolongation of survival only when recipients were IL-17(-/-) . Administration of a depleting anti-CD25 antibody to IL-17(-/-) recipients abrogated the survival advantage conferred by IL-17 deficiency, suggesting the involvement of a CD4(+) CD25(+) T cell regulatory mechanism. Therefore, IL-17 deficiency or neutralization was protective against the development of kidney allograft rejection, which may be mediated by impairment of Th1 responses and/or enhanced protection by Tregs.

The role of macrophages in the development of human renal allograft fibrosis in the first year after transplantation.

The aim of this study was to investigate the role of infiltrating macrophages in renal allograft fibrosis. Forty-six protocol renal allograft biopsies obtained 1 year after transplantation were stained with Sirius red to quantify fibrosis and double stained with CD68 and CD206 to identify the proportion of alternatively activated (M2) macrophages. Biopsies were analyzed for gene expression by microarray, which was correlated with macrophage infiltration and the severity of fibrosis. The number of infiltrating CD68+ cells strongly correlated with the percentage of interstitial fibrosis (r = 0.73, p < 0.0001). Macrophage infiltration at 1 year correlated with renal dysfunction at 1, 12 and 36 months posttransplant (estimated GFR low vs. high: 1 month 78 ± 26 vs. 54 ± 19 mL/min, p < 0.01; 12 months 87 ± 29 vs. 64 ± 19 mL/min, p < 0.05; 36 months 88 ± 33 vs. 60 ± 24 mL/min, p < 0.05). Ninety-two percent of infiltrating macrophages exhibited an M2 phenotype with CD68+ CD206+ dual staining. Gene microarrays demonstrated an alloimmune response with up-regulation of interferon-γ-response genes despite the lack of rejection or inflammatory infiltrate. Consistent with this was the presence of CXCL10 in proximal tubular cells at 3 months. This suggests that M2 macrophage proliferation, or infiltration, was associated with subclinical alloimmune inflammation, tubular injury and progression of fibrosis.

Pregnancy outcomes for kidney transplant recipients.

Pregnancy outcomes in a transplant population have not been well documented. Data from the Australia and New Zealand Dialysis and Transplant Registry (ANZDATA) and the National Perinatal Epidemiology and Statistics Unit (NPESU) were analyzed. We described pregnancy outcomes within the transplant population and compared these to outcomes for the general population. Six hundred ninety-two pregnancies in 447 transplant recipients were reported between 1971 and 2010 (ANZDATA); a corresponding 5 269 645 pregnancies were reported nationally in Australia between 1991 and 2010 (NPESU). At pregnancy transplant mothers had a median age of 31 years (interquartile range [IQR]: 27, 34), a median creatinine of 106 µmol/L (IQR: 88, 1103 µmol/L) and a functioning transplant for a median of 5 years (IQR: 3, 9). The mean gestational age at birth was 35 ± 5 weeks in transplant recipients, significantly shorter than the national average of 39 weeks (p < 0.0001). Mean live birth weight for transplant recipients was 873 g lower than the national average (2485 ± 783 g vs. 3358 ± 2 g); a significant difference remained after controlling for gestational age. There was lower perinatal survival rate in babies born to transplant recipients, 94% compared with 99% nationally (p < 0.001). Although transplant pregnancies are generally successful, outcomes differ from the general population, indicating these remain high-risk pregnancies despite good allograft function.

Infiltrating Foxp3(+) regulatory T cells from spontaneously tolerant kidney allografts demonstrate donor-specific tolerance.

Foxp3(+) regulatory T cells (Tregs) have an essential role in immune and allograft tolerance. However, in both kidney and liver transplantation in humans, FOXP3(+) Tregs have been associated with clinical rejection. Therefore, the role and function of graft infiltrating Tregs have been of great interest. In the studies outlined, we demonstrated that Foxp3(+) Tregs were expanded in tolerant kidney allografts and in draining lymph nodes in the DBA/2 (H-2(d) ) to C57BL/6 (H-2(b) ) mouse spontaneous kidney allograft tolerance model. Kidney allograft tolerance was abrogated after deletion of Foxp3(+) Tregs in DEpletion of REGulatory T cells (DEREG) mice. Kidney allograft infiltrating Foxp3(+) Tregs (K-Tregs) expressed elevated levels of TGF-ß, IL-10, interferon gamma (IFN-γ), the transcriptional repressor B lymphocyte-induced maturation protein-1 (Blimp-1) and chemokine receptor 3 (Cxcr3). These K-Tregs had the capacity to transfer dominant tolerance and demonstrate donor alloantigen-specific tolerance to skin allografts. This study demonstrated the crucial role, potency and specificity of graft infiltrating Foxp3(+) Tregs in the maintenance of spontaneously induced kidney allograft tolerance.

Selective depletion of alloreactive T cells leads to long-term islet allograft survival across a major histocompatibility complex mismatch in diabetic mice.

Islet cell transplantation as a therapy for type 1 diabetes has been limited by progressive graft loss. Significant immunosuppression including T-cell ablation has been used in an attempt to limit islet rejection. Here, we show that CD3(+) lymphocytes depleted of alloreactive T cells selected from a mixed lymphocyte reaction (MLR), where responder BALB/c splenocytes stained with carboxyfluorescein succinimidyl ester (CFSE) were stimulated with irradiated C57BL/6 splenocytes for 5 days, infused into diabetic immunodeficient mice are capable of restoring a broad T-cell repertoire and specifically do not reject islet transplants from the strain (C57BL/6) used in the original depletion. These mice demonstrate reconstitution with CD4(+) and CD8(+) T cells, the capacity to reject third-party grafts (CBA), and restoration of interferon-γ (IFN-γ) responses to third-party alloantigens. Over time, both forkhead box P3-positive (Foxp3(+)) T regulatory cells (Tregs) and γδ T cells expand, suggesting a role for peripheral tolerance, in addition to the initial depletion of alloreactive T cells, in long-term islet graft survival. Our results suggest that immune restoration with CD3(+) lymphocytes where alloreactive T cells are removed can restore cognate immunity without islet allograft loss and recurrence of diabetes.

CCL2 DNA vaccine to treat renal disease.

CCL2 DNA vaccines are directed against the host chemoattractant molecule CCL2 (MCP-1), a key chemokine in recruiting macrophages to sites of inflammation. Macrophages recruited by CCL2 lead to progressive renal injury. In rat models of disease unmodified CCL2 DNA vaccine in combination with a CCL5 (RANTES) DNA vaccine can protect against chronic renal disease. The mechanism of protection involves the induction of auto-antibodies to the CCL2. Introduction of the adjuvant p-tet into the DNA structure of the CCL2 vaccine leads to enhanced potency with the induction of specific Th1 cellular immunity. The strategies outlined here demonstrate a model for developing potent vaccines against highly restricted self targets.

Regulatory immune cells in kidney disease.

Lymphocytes and macrophages act as effector immune cells in the initiation and progression of renal injury. Recent data have shown that subpopulations of these immune cells (regulatory T lymphocytes and alternately-activated or regulatory macrophages) are potent modulators of tissue injury and repair in renal disease. Recent animal studies examining the therapeutic effect of these cells raise the exciting possibility that strategies targeting these cell types may be effective in treating and preventing kidney disease in humans. This review will describe their biological role in experimental kidney disease and therapeutic potential in clinical nephrology.

Ex vivo programmed macrophages ameliorate experimental chronic inflammatory renal disease.

Macrophage infiltration of the kidney is a prominent feature associated with the severity of renal injury and progressive renal failure. To determine the influence of macrophages in renal disease models in the absence of endogenous T and B cells, we performed adoptive transfer of macrophages into severe combined immunodeficient (SCID) mice. In this study, macrophages were isolated from the spleens of BALB/c mice and stimulated with lipopolysaccharide to induce classically activated M1 macrophages or with interleukin-4 (IL-4) and IL-13 to induce alternatively activated M2 macrophages. These macrophages were then infused into SCID mice with adriamycin nephropathy; an in vivo model of chronic inflammatory renal disease analogous to human focal segmental glomerulosclerosis. Mice infused with M1 macrophages had a more severe histological and functional injury, whereas M2 macrophage-induced transfused mice had reduced histological and functional injury. Both M1 and M2 macrophages localized preferentially to the area of injury and maintained their phenotypes even after 4 weeks. The protective effect of M2 macrophages was associated with reduced accumulation and possibly downregulated chemokine and inflammatory cytokine expression of the host infiltrating macrophages. Our findings demonstrate that macrophages not only act as effectors of immune injury but can be induced to provide protection against immune injury.

A protective role for programmed death 1 in progression of murine adriamycin nephropathy.

Programmed death 1 (PD-1) is a novel member of the CD28/cytotoxic T-lymphocyte-associated protein-4 superfamily, which plays an important role in the regulation of activated T cells. However, it is not clear how PD-1 is expressed in normal and diseased kidney, nor if it has a role in progression of chronic renal disease. PD-1 expression and the effect of monoclonal anti-PD-1 antibody (Ab) were examined in murine adriamycin nephropathy (AN). BALB/c mice were divided into three groups: (a) normal mice, (b) adriamycin (ADR) with control immunoglobulin (Ig)G (ADR-IgG), and (c) ADR with anti-PD-1 Ab (ADR-Ab). AN was induced by a single intravenous injection of ADR. Anti-PD-1 Ab was given by intraperitoneal injection on alternate days from day 0 to day 10, or to day 18. Animals were killed at week 4. Renal function, histological change, and cytokine expression were examined. PD-1 mRNA was detected in kidney tissue of mice with AN in a dose- and time-dependent manner. PD-1 was mainly expressed on injured tubule cells and some interstitial cells, which co-stained with alpha-smooth muscle actin in AN, but not in normal kidney. Anti-PD-1 treatment up to day 18, but not to day 10, worsened glomerular and tubulointerstitial injury. The ratio of urinary protein/creatinine was significantly higher in ADR-Ab mice than ADR-IgG mice. The number of macrophages was significantly increased in ADR-Ab mice compared with ADR-IgG mice. Blockade of PD-1 worsened progressive renal histopathological and functional injury in murine AN. This suggests a possible protective role for PD-1 in chronic renal disease, and its potential as a treatment to slow disease progression.

NK cells do not mediate renal injury in murine adriamycin nephropathy.

In adriamycin nephropathy (AN), a model of chronic proteinuric renal injury, the absence of functional B and T cells with residual natural killer (NK) cells, and macrophages in severe combined immunodeficient (SCID) mice results in more severe disease than in immunocompetent mice. We have recently shown expression of the stimulatory NK cell molecule NKG2D and its ligand RAE-1 in the adriamycin (ADR) kidney. Therefore, we sought to determine the role of NK cells in AN. We used anti-asialo GM1 NK cell depletion in immunocompetent BALB/c mice with AN, and also compared AN in immunodeficient SCID mice and immunodeficient nonobese diabetic (NOD)-SCID mice (that have impaired NK cell function). The number of NK cells was increased in AN in BALB/c mice compared with normal controls. NK cell depletion or reduction of NK function in NOD-SCID mice did not affect the severity of disease. In both wild type and immunodeficient models, ADR upregulated RAE-1 in the kidney. High levels of Class I major histocompatibility complex molecules were found in both models of AN. In conclusion, NK cells do not play a significant role in AN.

Adenovirus associated haematuria.

Adenovirus is a common respiratory virus in children and is known to cause acute haemorrhagic cystitis, particularly in the immunosuppressed. In immunocompetent children with adenoviral infection the incidence of haematuria was 18.6%, with 2.4% of these children having macroscopic haematuria and upper tract involvement.

IL-12, STAT4-dependent up-regulation of CD4(+) T cell core 2 beta-1,6-n-acetylglucosaminyltransferase, an enzyme essential for biosynthesis of P-selectin ligands.

TCR activation of naive T cells in the presence of IL-12 drives polarization toward a Th1 phenotype and synthesis of P- and E-selectin ligands. Fucosyltransferase VII (Fuc-T VII) and core 2 beta-1,6-N-acetylglucosaminyltransferase (C2GnT) are critical for biosynthesis of selectin ligands. P-selectin glycoprotein ligand-1 is the best characterized ligand for P-selectin and also binds E-selectin. The contributions of TCR and cytokine signaling pathways to up-regulate Fuc-T VII and C2GnT during biosynthesis of E- and P-selectin ligands, such as P-selectin glycoprotein ligand 1, are unknown. IL-12 signals via the STAT4 pathway. Here, naive DO11.10 TCR transgenic and STAT4(-/-) TCR transgenic CD4(+) T cells were stimulated with Ag and IL-12 (Th1 condition), IL-4 (Th2), or neutralizing anti-IL-4 mAb only (Th0). The levels of Fuc-T VII and C2GnT mRNA in these cells were compared with their adhesive interactions with P- and E-selectin in vitro under flow. The data show IL-12/STAT4 signaling is necessary for induction of C2GnT, but not Fuc-TVII mRNA, and that STAT4(-/-) Th1 cells do not traffic normally to sites of inflammation in vivo, do not interact with P-selectin, and exhibit a partial reduction of E-selectin interactions under shear stress in vitro. Ag-specific TCR activation in CD4(+) T cells was sufficient to trigger induction of Fuc-TVII, but not C2GnT, mRNA and expression of E-selectin, but not P-selectin, ligands. Thus, Fuc-T VII and C2GnT are regulated by different signals during Th cell differentiation, and both cytokine and TCR signals are necessary for the expression of E- and P-selectin ligands.

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.

Regulatory interactions of alphabeta and gammadelta T cells in glomerulonephritis.

BACKGROUND: Several lines of evidence suggest that cellular immune mechanisms contribute to glomerulonephritis. METHODS: The roles of alphabeta and gammadelta T cells in the pathogenesis of glomerulonephritis were investigated in a model of nephrotoxic nephritis in mice deficient in either T-cell population [T-cell receptor (TCR)beta and TCRdelta knockout mice]. The model, induced by the injection of rabbit anti-mouse glomerular basement membrane antibody, is characterized by the development of proteinuria and glomerular damage over a 21-day observation period in wild-type mice. RESULTS: Mice deficient in either alphabeta or gammadelta T cells developed minimal proteinuria and glomerular lesions and had a significant reduction in macrophage accumulation compared with wild-type mice. In gammadelta T-cell-deficient mice, circulating levels and glomerular deposition of autologous IgG were comparable to wild-type levels, while alphabeta T-cell-deficient mice had no autologous IgG production. Autologous antibody production was not required for the development of glomerulonephritis since mice that lack IgG and B cells (micro-chain-/-) developed similar proteinuria to that observed in wild-type mice. CONCLUSIONS: These studies suggest a proinflammatory role for both alphabeta and gammadelta T cells in glomerular injury, independent of the humoral response. This is the first demonstration, to our knowledge, that both T-cell subsets contribute to the progression of a disease, and it suggests that complex regulatory interactions between alphabeta and gammadelta T cells play a role in glomerular injury.

Expression of B7 molecules in recipient, not donor, mice determines the survival of cardiac allografts.

Blockade of the CD28/CTLA4/B7 costimulatory pathway using CTLA4-Ig has great therapeutic potential, and has been shown to prolong allograft survival in a variety of animal models. To gain further insight into the mechanism by which costimulatory blockade prevents allograft rejection, we studied cardiac allograft survival in the complete absence of B7 costimulation using mice lacking B7-1 and B7-2 (B7-1/B7-2-/- mice). To determine the role of B7 on donor vs recipient cells, we used B7-1/B7-2-/- mice as either donors or recipients of allografts. Wild-type (WT) recipients acutely reject fully allogeneic hearts from both WT and B7-1/B7-2-/- mice. In contrast, B7-1/B7-2-/- recipients allow long-term survival of grafts from both WT and B7-1/B7-2-/- mice, with minimal histologic evidence of either acute or chronic rejection in grafts harvested after 90 days. The B7-1/B7-2-/- mice acutely reject B7-1/B7-2-/- allografts if CD28 stimulation is restored by the administration of Ab to CD28 and can mount an alloresponse in mixed lymphocyte reactions. Therefore, B7-1/B7-2-/- mice are capable of generating alloresponses both in vivo and in vitro. Our results demonstrate that in the alloresponse to mouse heterotopic cardiac transplantation, B7 molecules on recipient cells rather than donor cells provide the critical costimulatory signals. The indefinite survival of allografts into B7-1/B7-2-/- recipients further shows that the absence of B7 costimulation alone is sufficient to prevent rejection.

Endothelial cells modify the costimulatory capacity of transmigrating leukocytes and promote CD28-mediated CD4(+) T cell alloactivation.

Activated vascular endothelial cells (ECs) express major histocompatibility complex (MHC) class II molecules in vitro and in vivo in acute and chronic allograft rejection. However, human ECs may be limited in their ability to effectively activate CD4(+) T cells, because they do not express members of the B7 family (CD80 and CD86) of costimulatory molecules. In this study, we show that ECs promote the full activation of CD4(+) T cells via trans-costimulatory interactions. By reverse transcriptase polymerase chain reaction, Western blot, and FACS((R)) analysis, we could not detect the expression of CD80 and CD86 on activated ECs and found minimal expression on purified CD4(+) T cells. In contrast, both CD80 and CD86 were expressed in allogeneic CD4(+) T cell-EC cocultures. Expression of CD86 peaked at early times between 12 and 24 h after coculture, whereas CD80 was not expressed until 72 h. Addition of anti-CD86 but not anti-CD80 monoclonal antibodies to cocultures inhibited IL-2 production and the proliferation of CD4(+) T cells to allogeneic donor human umbilical vein ECs (HUVECs), as well as to skin and lung microvascular ECs. Furthermore, we found that interferon gamma-activated ECs but not untreated ECs induced mRNA and cell surface expression of CD80 and CD86 on CD4(+) T cells, and these T cells were functional to provide a trans-costimulatory signal to autologous CD4(+) T cells. Blockade of MHC class II and lymphocyte function-associated antigen 3 but not other EC cell surface molecules on IFN-gamma-activated ECs inhibited the induction of CD86 on CD4(+) T cells. Transmigration of purified populations of monocytes across EC monolayers similarly resulted in the induction of functional CD86, but also induced the de novo expression of the cytokines interleukin (IL)-1alpha and IL-12. In addition, EC-modified monocytes supported enhanced proliferation of allogeneic and autologous CD4(+) T cells. Taken together, these data define the ability of the endothelium to modify CD4(+) T cells and monocytes for trans-costimulatory events. This unique function of the endothelium in alloimmune T cell activation has functional consequences for the direct and the indirect pathways of allorecognition.

Inhibition of allorecognition by a human class II MHC-derived peptide through the induction of apoptosis.

The interaction of the T-cell receptor with the major histocomatibility complex (MHC)-peptide complex is central to T-cell activation. Variation in the nature of the peptide bound within the groove of the MHC molecule may result in an altered T-cell response. Because some naturally processed peptides bound within the groove of the class II MHC molecule are derived from the MHC molecules themselves, we studied the inhibitory effects of synthetic class II MHC peptides on alloimmune responses in vitro. Three peptides derived from a highly conserved region of the class II MHC alpha chains inhibited the rat mixed lymphocyte response (MLR) in a dose-dependent manner, with the human HLA-DQA1 peptide also inhibiting the human and mouse MLR. No effect was seen on mitogen-induced T-cell proliferation. HLA-DQA1 inhibited cytolytic T lymphocyte (CTL) generation in a dose-response fashion, with no reduction in preformed CTL killing, suggesting that the inhibitory effect is targeted at CD4(+) T-cell function. Cell-cycle analysis by flow cytometry showed that restimulation of primed T cells in the presence of HLA-DQA1 resulted in increased apoptosis, whereas unstimulated cells were not affected. These data demonstrate that synthetic peptides derived from highly conserved regions of the class II MHC alpha chain can alter CD4(+) T-lymphocyte alloimmune responses in vitro, and this effect is mediated by the induction of apoptosis in activated T cells.