Peptide length-dependent TCR antagonism on class II HLA-restricted responses of peripheral blood mononuclear cells and T cell clones

T cell clonal recognition of peptide ligands is highly diverse. To investigate how peptide mixtures with diverse sequences affect polyclonal responses of peripheral blood mononuclear cells (PBMC), we synthesized Xn (n = 9-19) peptides that consist of 9 to 19 residues with random sequences. We found that: (1) in antagonism assays, Xn peptides inhibit polyclonal responses of PBMC induced by purified protein derivative (PPD) and crude mite extracts as well as mixed lymphocyte reaction (MLR), in which intermediate-length peptides (n = 13 or 15) show the largest inhibitory effects; and (2) a high-affinity HLA-DR4-binding peptide is devoid of inhibitory activity against MLR to DR4, indicating that these effects are not caused by inhibition of peptide binding to HLA. Furthermore, X15 did not abrogate PBMC proliferation induced by phorbol 12-myristate 13-acetate + ionomycin, or anergize PBMC by preculture. All these observations indicate that TCR antagonism does exist at peripheral T cell levels in humans, and that Xn peptides, depending on peptide length, are capable of antagonizing T cell polyclonal responses. Indeed, even with cloned T cells, certain non-agonistic peptides shorter (but not too short) than the wild type in their C termini, function as TCR antagonists, findings which corroborate the observation that X13-15 antagonizes T cell responses more efficiently than does X17-19 or X9-11.     

Minimal peptide length requirements for CD4(+) T cell clones--implications for molecular mimicry and T cell survival

CD4(+) T lymphocytes usually recognize peptides of 12-16 amino acids in the context of HLA class II molecules. We have recently used synthetic peptide combinatorial libraries to dissect in detail antigen recognition by autoreactive CD4(+) T cell clones (TCC). The results of these studies demonstrated that antigen recognition by T cells is highly degenerate and that many cross-reactive ligands can be defined, some of which much more potent than the selecting autoantigen. Based on these observations, we examined the response of a myelin basic protein-specific HLA class II-restricted CD4(+) TCC to truncation variants of optimal ligands. Surprisingly, pentapeptides, tetrapeptides and even tripeptides derived from different segments of the optimal ligands were recognized by the TCC, and some were even more potent than the selecting autoantigen. In addition, these peptides enhanced the survival of the TCC at low concentration. The relevance of this finding was supported by the generation of pentapeptide-specific CD4(+) TCC from peripheral blood lymphocytes. These observations not only change existing views on the length requirements for activation of CD4(+) HLA class II-restricted T cells, but also extend our knowledge about the flexibility of TCR recognition and the potential for cross-reactivity in the immune system.     

Discrimination of human CD4 T cell clones based on their reactivity with antigen-presenting T cells.

In this report, we describe the discrimination of human T cell clones based on their reactivity with activated T cells as antigen-presenting cells (APC). CD4+ T cell clones specific for peptide P30 of tetanus toxin (amino acids 947-967) and restricted to the DP4 molecule were established and tested for proliferation to peptide presented either by peripheral blood mononuclear cells (PBMC), Epstein-Barr virus (EBV)-transformed B cells or major histocompatibility complex (MHC) class II-expressing T cells. We found two sets of T cell clones: one set proliferated to peptide presentation by PBMC, EBV-transformed B cell lines (EBV-B cells) and MHC class II+ T cells (termed T-responder clones), while the other set of clones was only stimulated to proliferate, if the peptide was presented by PBMC or EBV-B cells, but not by T cells (T-nonresponder clones). Nevertheless, these T-nonresponder clones recognized P30 also on T cells, as revealed by Ca2+ influx. The discrimination of the clones was not due to different avidities of the T cell receptors (TcR) of individual clones for the MHC-peptide complex as T-responder and T-nonresponder clones had similar dose-response curves to P30 presented by fixed EBV-B cell lines. Addition of cytokines [interleukin (IL)-1, IL-2, IL-4 and interferon gamma] did not change the proliferative response of the clones, which was consistent throughout an observation period of greater than 4 months. T-nonresponder clones, exposed to P30 on MHC class II-expressing T cells, became not anergic, as they could be restimulated by P30 presented on EBV-B cells. The measurement of a panel of T cell activation markers and adhesion molecules on T-responder and T-nonresponder clones revealed a higher expression of the CD28 molecule on the T-nonresponder clones. The data suggest that freshly cloned T cells can be differentiated by peptide presentation on classical (PBMC, EBV-B cells) or non-classical APC (class II+ T cells), and that this discrimination is further underlined by different levels of adhesion molecules.     

Soluble CD14 acts as a negative regulator of human T cell activation and function

T cell activation is controlled by the coordination of stimulatory and negative regulatory signals which are not completely defined. In this study we tested for a possible direct effect of CD14 on the regulation of T cell activation and function. We show that soluble CD14 (sCD14) induces inhibition of antigen-mediated peripheral blood mononuclear cells (PBMC) proliferation and anti-CD3-mediated proliferation of CD4⁺CD8⁻, CD4⁻CD8⁺ and CD4⁺CD8⁺ T cell clones. This effect is not due to cell death, but results from a marked inhibition of IL-2 production. Proliferation of T cell clones due to exogenous IL-2 is not affected by sCD14. We also found that sCD14 inhibits production of another Th1-like cytokine, IFN-γ and a Th2-like cytokine, IL-4. Importantly, sCD14 induces a progressive accumulation of the inhibitory protein IκB-α. We show that sCD14 binds to activated T cells. Following cell activation, biotinylated sCD14 stains CD3⁺ PBMC, as well as human T cell clones with varying intensity. The binding is saturable, can be inhibited by excess of unlabeled sCD14 and, following binding, sCD14 is internalized. Collectively, these findings reveal a previously unrecognized function of sCD14, namely its capacity to negatively regulate T lymphocyte activation and function by interacting directly with activated T cells.     

Decreased specific CD8+ T cell cross-reactivity of antigen recognition following vaccination with Melan-A peptide

The aim of T cell vaccines is the expansion of antigen-specific T cells able to confer immune protection against pathogens or tumors. Although increase in absolute cell numbers, effector functions and TCR repertoire of vaccine-induced T cells are often evaluated, their reactivity for the cognate antigen versus their cross-reactive potential is rarely considered. In fact, little information is available regarding the influence of vaccines on T cell fine specificity of antigen recognition despite the impact that this feature may have in protective immunity. To shed light on the cross-reactive potential of vaccine-induced cells, we analyzed the reactivity of CD8(+) T cells following vaccination of HLA-A2(+) melanoma patients with Melan-A peptide, incomplete Freund's adjuvant and CpG-oligodeoxynucleotide adjuvant, which was shown to induce strong expansion of Melan-A-reactive CD8(+) T cells in vivo. A collection of predicted Melan-A cross-reactive peptides, identified from a combinatorial peptide library, was used to probe functional antigen recognition of PBMC ex vivo and Melan-A-reactive CD8(+) T cell clones. While Melan-A-reactive CD8(+) T cells prior to vaccination are usually constituted of widely cross-reactive naive cells, we show that peptide vaccination resulted in expansion of memory T cells displaying a reactivity predominantly restricted to the antigen of interest. Importantly, these cells are tumor-reactive.     

Engagement of IL-1 receptor accessory protein (IL-1RAcP) with the monoclonal antibody AY19 provides co-activating signals and prolongs the CD2-induced proliferation of peripheral blood lymphocytes

IL-1 receptor accessory protein (IL-1RAcP) is the second subunit required to form a functional receptor complex for IL-1α and β, IL-1F6, IL-1F8, IL1-F9 and IL-33. While it does not directly interact with the cytokines, IL-1RAcP is necessary to mediate signal transduction. We previously reported a monoclonal antibody with an unknown specificity, termed AY19, that was capable to induce a significant increase in the size of CFU-GM colonies when added to cultures of human cord blood CD34(+) hematopoietic progenitors. Here we demonstrate that AY19 mAb recognizes IL1-RAcP. We show that this adaptor molecule is significantly present on peripheral blood monocytes and lymphocytes including CD4(+) and CD8(+) T lymphocytes, B and NK cells. Interestingly, its expression is found increased on CD127(low)CD4(+)CD25(high) T cells when compared to CD127(low)CD4(+)CD25(-) T cell subset, suggesting that the level of IL-1RAcP membrane expression could allow to distinguish within CD127(low)CD4(+) T lymphocytes the CD25(high) T regulatory subset from conventional CD25(-) T lymphocytes. Functional studies reveal that addition of AY19 mAb enhances the proliferation of peripheral blood mononuclear cells (PBMC) obtained with mitogenic concentrations of PMA. Interestingly, we found that although AY19 mAb does not increase the optimal PBMC proliferation induced by a mitogenic pair of anti-CD2 mAbs it prolongs their time of proliferation. Thus, these results indicate that the anti-IL-1RAcP mAb AY19 exhibits unique functional properties by triggering co-stimulatory signals in lymphocytes.     

The polyomavirus BK large T-antigen-derived peptide elicits an HLA-DR promiscuous and polyfunctional CD4+ T-cell response

BK virus (BKV) nephropathy and hemorrhagic cystitis are increasingly recognized causes of disease in renal and hematopoietic stem cell transplant recipients, respectively. Functional characterization of the immune response to BKV is important for clinical diagnosis, prognosis, and vaccine design. A peptide mix (PepMix) and overlapping (OPP) or random (RPP) peptide pools derived from BKV large T antigen (LTA) were used to restimulate 14-day-expanded peripheral blood mononuclear cells (PBMC) from 27 healthy control subjects in gamma interferon (IFN-γ)-specific enzyme-linked immunospot (ELISPOT) assays. A T-cell response to LTA PepMix was detected in 15/27 subjects. A response was frequently observed with peptides derived from the helicase domain (9/15 subjects), while the DNA binding and host range domains were immunologically inert (0/15 subjects). For all nine subjects who responded to LTA peptide pools, the immune response could be explained largely by a 15-mer peptide designated P313. P313-specific CD4(+) T-cell clones demonstrated (i) stringent LTA peptide specificity; (ii) promiscuous recognition in the context of HLA-DR alleles; (iii) cross recognition of homologous peptides from the polyomavirus simian virus 40 (SV40); (iv) an effector memory phenotype, CD107a expression, and intracellular production of IFN-γ and tumor necrosis factor alpha (TNF-α); (v) cytotoxic activity in a chromium release assay; and (vi) the ability to directly present cognate antigen to autologous T cells. In conclusion, T-cell-mediated immunity to BKV in healthy subjects is associated with a polyfunctional population of CD4(+) T cells with dual T-helper and T-cytotoxic properties. HLA class II promiscuity in antigen presentation makes the targeted LTA peptide sequence a suitable candidate for inclusion in immunotherapy protocols.     

Evidence for shared recognition of a peptide ligand by a diverse panel of non-obese diabetic mice-derived,islet-specific,diabetogenic T cell clones

MHC class II-restricted autoreactive T cells play a major role in the development of autoimmune diabetes mellitus in both human and mouse. Two of our groups previously established panels of islet-reactive CD4+ T cell clones from prediabetic non-obese diabetic (NOD) mice. These clones express distinct sets of TCR V alpha , V beta , J alpha and J beta , and also differ in the structure of the junctional region of TCR. All of the T cell clones have been shown to cause insulitis and several induce diabetes when transferred to various recipients. The antigen specificities of these T cell clones have not been determined, but they do not react with defined islet cell antigens such as glutamic acid decarboxylase. To identify the peptide ligands recognized by these clones, we examined the reactivity of the T cell clones to peptide mixtures in which anchor residues for H2-A g7 were fixed. Most of the clones showed similar reactivity to the peptide mixtures. To further determine the peptide ligands of the T cell clones, we synthesized several peptides based on the favored amino acid motifs and examined clone reactivity to the synthetic peptides. Some of the peptides, e.g. HLAI-RM and HIPI-RM, could stimulate most of the T cell clones tested, even though the clones expressed different TCR. The results suggest that our islet-reactive T cell clones recognize in islet beta cells a natural ligand that is similar to these peptides.     

Therapeutic effect of an altered peptide ligand derived from heat-shock protein 60 by suppressing of inflammatory cytokines secretion in two animal models of rheumatoid arthritis

Rheumatoid arthritis is a systemic autoinmmune disease mediated by T cells. Productive engagement of T cell receptors by major histocompatibility complex-peptide leads to proliferation, differentiation and the definition of effector functions. Altered peptide ligands (APL) generated by amino acid substitutions in the antigenic peptide have diverse effects on T cell response. We predicted a novel T cell epitope from human heat-shock protein 60, an autoantigen involved in the pathogenesis of rheumatoid arthritis. Three APLs were designed from this epitope and it was demostrated that these peptides induce the activation of T cells through their ability to modify cell cycle phase's distribution of CD4+T cells from RA patients. Also, IL-17, TNF-α and IL-10 levels were determined in PBMC from these patients. Unlike the wild-type peptide and the other two APLs, APL2 increased the IL-10 level and suppressed IL-17 secretion in these assays. Therapeutic effect of this APL in adjuvant arthritis (AA) and collagen-induced arthritis (CIA) models was also evaluated. Clinical score, histopathology, inflammatory and regulatory cytokine concentration were monitored in the animals. APL2 efficiently inhibited the progression of AA and CIA with a significant reduction of the clinical and histopathogic score. Therapeutic effect of APL2 on CIA was similar to that obtained with MTX; the standard treatment for RA. This effect was associated with a decrease of TNF-α and IL-17 levels. These results suggest that the therapeutic effect of APL2 is mediated in part by down-regulation of inflammatory cytokines and support the potential use of APL2 as a therapeutic drug in RA patients.     

Regulation of cytokine production in human peripheral blood mononuclear cells and allergen-specific th cell clones by 1alpha,25-dihydroxyvitamin D3

BACKGROUND: The steroid hormone 1alpha,25-dihydroxyvitamin D3 (calcitriol), in addition to its crucial role in calcium homeostasis, exerts several effects on the immune system by regulating cell proliferation, differentiation, and maturation. These effects may be exerted through the control of protooncogenes and the regulation of cytokine production. METHODS: The influence of calcitriol on cytokines secretion by human peripheral blood mononuclear cells (PBMC) isolated from healthy donors, and by allergen-specific T helper (Th) cell clones was studied. PBMC were cultured for 48 h with phorbol myristate acetate (PMA) and ionomycin in the presence or absence of calcitriol. Human Th cell clones were stimulated with either Bet v 1 allergen or anti-CD3 antibodies and PMA. Cytokines were measured in the supernatants by ELISA, and at single-cell level by FACS. RESULTS: Calcitriol significantly inhibited the production of IL-2, IFN-gamma and IL-12 by PBMC, as well as the percentage of CD4+ T cells containing intracytoplasmic IL-2 and IFN-gamma. Interestingly, calcitriol-treated PBMC induced the production of IL-10 and IL-5, but not of IL-4. The effect of calcitriol was maximal at 10(-7) to 10(-9) and noneffective at 10(-11) M. Calcitriol diminished the secretion of IL-1, TNF-alpha, and MG-CSF in PBMC. Furthermore, calcitriol also decreased the secretion of IL-2 and IFN-gamma by Th1 clones, and of IL-4 by Th2 clones. CONCLUSIONS: Our data strongly support the notion that calcitriol modulates the production of cytokines in a time- and concentration-dependent manner, and suggest that nonhypercalcemic derivatives of 1alpha,25-dihydroxyvitamin D(3) may be used for new immunosuppressive therapies.     

不同IL-15基因转染对NCI-H446细胞诱导PBMC增殖和杀伤肿瘤细胞的影响

目的 研究不同IL-15基因转染对NCI-H446细胞诱导外周血单个核细胞(PBMC)增殖和杀伤肿瘤细胞的影响.方法 野生型NCI-H446细胞(Cw)和被3种IL-15基因分别转染的3种NCI-H446细胞(Cmp:被IL-15成熟肽基因转染;Cp:被原型IL-15基因转染;Csp:被信号肽换为IL-2信号肽的改型IL-15基因转染),用丝裂霉素(M)处理后(分别名为MCw、MCmp、MCp和MCsp)作为刺激细胞刺激健康志愿者的PBMC.对这些被刺激的PBMC,分别名为MCw-PBMC、MCmp-PBMC、MCp-PBMC和MC-sp-PBMC.台盼蓝拒染法计数细胞数,流式细胞术测定CD4 细胞和CD8 细胞百分率.在MCmp-PBMC,MCp-PBMC和MCsp-PBMC中,选择其细胞数和CD4 细胞和/或CD8 细胞百分率统计学上明显高于MCw-PBMC的作为效应细胞,MTT法测定它们对Cw的杀伤.结果 与MCw-PBMC相比,MCp-PBMC在细胞数、CD4 细胞和CD8 细胞百分率及对Cw的杀伤上,均显著提高(P＜0.05).结论 原型IL-15基因转染能提高NCI-H446细胞诱导PBMC增殖和杀伤野生型NCI-H446细胞的能力.     

BCG induced CD4+ cytotoxic T cells from BCG vaccinated healthy subjects: relation between cytotoxicity and suppression in vitro.

Mycobacterial antigen specific cytotoxic T cells killing antigen-pulsed antigen presenting cells (APC) were induced from peripheral blood mononuclear cells (PBMC) of BCG-vaccinated healthy subjects after activation in vitro with BCG. CD8+ depleted cells were as effective as PBMC, indicating that CD4+ cells play a dominant role in this phenomenon. CD4+ T cell clones raised against BCG also exhibited mycobacterial antigen specific cytotoxicity and suppressed BCG-driven selfproliferation. However, the same clones could either suppress or enhance the proliferation of other T cell clones from the same subject. The possible function in vivo of the cytotoxicity mediated by CD4+ T cells is discussed.     

Differential peptide binding to CD40 evokes counteractive responses

The antigen-presenting cell–expressed CD40 is implied in the regulation of counteractive immune responses such as induction of pro-inflammatory and anti-inflammatory cytokines interleukin (IL)–12 and IL-10, respectively. The mechanism of this duality in CD40 function remains unknown. Here, we investigated whether such duality depends on ligand binding. Based on CD40 binding, we identifed two dodecameric peptides, peptide-7 and peptide-19, from the phage peptide library. Peptide-7 induces IL-10 and increases Leishmania donovani infection in macrophages, whereas peptide-19 induces IL-12 and reduces L. donovani infection. CD40-peptide interaction analyses by surface plasmon resonance and atomic force microscopy suggest that the functional differences are not associated with the studied interaction parameters. The molecular dynamic simulation of the CD40-peptides interaction suggests that these two peptides bind to two different places on CD40. Thus, we suggest for the first time that differential binding of the ligands imparts functional duality to CD40.     

Rescue of memory CD8+ T cell reactivity in peptide/TLR9 ligand immunization by codelivery of cytokines or CD40 ligation

The capability of cellular immune components to rapidly recall upon challenge in most situations decides the efficacy of a vaccine. Here, we show that immunization of mice with SSIEFARL peptide (immunodominant epitope in glycoprotein B of herpes simplex virus type 1, aa498-505) combined with TLR9 ligand in the absence of helper CD4(+) T cell activation generates a functionally impaired CD8(+) T cell memory response. Codelivery of IL-12, IL-15, or anti-CD40 together with MHC class-I-restricted peptide combined with TLR9 ligand at inception of immunization resulted in generation of memory CD8(+) T cells that were several fold less compromised than immunization with peptide alone. Furthermore, administration of either plasmid DNA encoding IL-15 or anti-CD40 mAb but not rIL-12 during the memory phase restored the reactivity of memory CD8(+) T cells. Moreover, the rescued CD8(+) T cells preserved their cytotoxic capability and were able to clear a recombinant vaccinia virus encoding glycoprotein B of HSV. Our results indicate that good memory CD8(+) T cell response to peptide immunization can be achieved by using costimulatory procedures at the time of priming or recall immunization.     

Regulation of interleukin-4 production in human mononuclear cells.

Interleukin (IL)-4 is a cytokine with a broad range of effects on immune cells, however, little is known regarding the regulation of its production in freshly isolated human peripheral blood mononuclear cells (PBMC). Here we report the production of IL-4 in such cells following stimulation with monoclonal antibodies (mAb) directed against different cell surface antigens. We show that triggering via CD2 is more efficient for IL-4 production than triggering via the CD3 complex. The addition of a CD28 mAb enhances IL-4 production approximately threefold. Cell depletion experiments show that among CD2 plus CD28-stimulated PBMC the production of IL-4 is restricted to the CD8-CD45RA-T cell subpopulation. mAb interfering with the binding of IL-2 to its receptor can inhibit the production of IL-4 in CD2 plus CD28-stimulated PBMC. As IL-2 induces cell proliferation and production of interferon-gamma, but not production of IL-4, it follows that IL-2 is necessary but not sufficient for IL-4 production.     

Interferon-beta induces selective enhancement of antigen-specific T cell responses.

Interferon-beta (IFN-beta) inhibits mitogen-induced T cell responses, in part through downregulation of interleukin-12 (IL-12) or upregulation of IL-10. We have reexamined these findings using ragweed (RW) stimulated or tetanus toxoid (TT)-stimulated human peripheral blood mononuclear cells (PBMC) and nontransformed, antigen-specific, human Th0, Th1, and Th2 clones. IFN-beta induced concentration-dependent inhibition of phytohemagglutinin (PHA)-stimulated PBMC proliferation and enhancement of RW-stimulated or TTstimulated PBMC proliferation. Monocyte depletion of PBMC isolates resulted in concentration-dependent inhibition of RW-driven or TT-driven proliferation by IFN-beta. This response was unaltered by the addition of either exogenous recombinant human IL-12 (rHuIL-12) or saturating concentrations of anti-IL-10. Moreover, addition of exogenous rHuIL-10 to nondepleted RW-driven or TT-driven PBMC cultures did not alter the concentration-dependent enhancement of antigen-driven proliferation induced by IFN-beta. Th0, Th1, and Th2 clones stimulated in the presence of antigen and autologous, irradiated PBMC displayed concentration-dependent inhibition of proliferation in the presence of IFN-beta that was unaltered by the addition of either exogenous rHuIL-12 or a saturating concentration of anti-IL-10. Finally, whereas IFN-beta inhibited antigen-driven generation of IL-5, IL-12, IL-13, and IFN-gamma, IFN-beta enhanced generation of both IL-4 and IL-10. Thus, IFN-beta, induces a selective, IL-10-independent and IL-12-independent upregulation of antigen-specific T cell responses, supporting the role of IFN-beta as an immunomodulatory rather than an antiproliferative/immunosuppressive cytokine.     

Identification of broadly recognized, T helper 1 lymphocyte epitopes in an equine lentivirus

Equine infectious anaemia virus (EIAV) is a horse lentivirus causing lifelong, persistent infection. During acute infection, CD8(+) cytotoxic T lymphocytes (CTL) are probably involved in terminating plasma viraemia. However, only a few EIAV CTL epitopes, restricted to fewer horse major histocompatibility complex (MHC) class I alleles, are known. As interferon-gamma (IFN-gamma)-secreting CD4(+), T helper 1 (Th1) lymphocytes promote CTL activity and help maintain memory CTL, identifying broadly recognized EIAV Th1 epitopes would contribute significantly to vaccine strategies seeking to promote strong CTL responses among horses with varying class I haplotypes. To this end, peripheral blood mononuclear cells (PBMC) from 10 MHC disparate, EIAV-infected horses were tested in T-lymphocyte proliferation assays for recognition of peptides from the Gag p26 capsid region and a portion of Pol. Both regions are highly conserved among EIAV isolates, and this Pol region is 51-63% homologous to other lentiviral Pol proteins. Seven of 10 horses recognized peptide Gag 221-245, and peptides Gag 242-261 and Pol 323-344 were recognized by five and four horses, respectively. Furthermore, the Gag peptides were recognized by two additional horses after resolving their initial plasma viraemia, indicating that these two peptides can be immunodominant early in infection. Gag peptide-responsive PBMC produced only IFN-gamma, indicating a Th1 response, while Pol 323-344-responsive PBMC produced IFN-gamma both with and without interleukin-4. PBMC from uninfected horses failed to either proliferate or secrete cytokines in response to peptide stimulation. Finally, CD4(+) T lymphocytes were required for proliferation responses, as shown by assays using CD4- versus CD8-depleted PBMC.     

Cyclosporin A inhibition of macrophage colony-stimulating factor (M-CSF) production by activated human T lymphocytes

M-CSF is a pleiotropic cytokine involved in the survival, proliferation, and differentiation of cells of the monocyte/macrophage lineage. M-CSF is produced by numerous cells including CD3-activated T cells. M-CSF serum levels are increased during acute graft rejection. We tested the in vitro production of M-CSF, GM-CSF, IL-2, and IL-4 by T-cell clones costimulated by CD3 and accessory activation pathways and the effects of cyclosporin A and methylprednisolone. The nine clones studied and CD4+ cells purified from peripheral blood mononuclear cells (PBMC) spontaneously produced low levels of M-CSF, which PMA and CD3 mAb strongly enhanced. In contrast to IL-2, CD28 mAb did not further enhance this production. CsA inhibited M-CSF production by clones and purified CD4 T cells. Addition of IL-2, anti IL-2, or anti CD25 mAb to the cultures demonstrated that CsA down-regulated M-CSF synthesis by activated T cells through its inhibition of IL-2 synthesis. These results could help to better understand the complex mechanisms of acute graft rejection and immunosuppression.     

胚胎骨髓来源间充质干细胞对人Th17细胞的免疫调节

背景：众多研究表明间充质干细胞能发挥免疫调节功能,抑制T细胞增殖。 目的：观察胚胎骨髓来源间充质干细胞对人Th17细胞的调节作用。 方法：将人胚胎骨髓间充质干细胞与正常人外周血单个核细胞或CD4⁺ T细胞以1∶10比例共培养4 d,以单个核细胞或CD4⁺T细胞单独培养为对照。应用实时定量PCR检测细胞白细胞介素17 mRNA表达,酶联免疫吸附试验检测细胞上清中白细胞介素17蛋白水平,流式细胞术检测Th17细胞数量。 结果与结论：胚胎骨髓来源间充质干细胞与单个核细胞共培养组白细胞介素17 mRNA表达水平明显高于单个核细胞组(P < 0.01)。与此一致的是,胚胎骨髓来源间充质干细胞与单个核细胞或CD4⁺T细胞共培养组细胞上清中白细胞介素17蛋白水平明显高于单个核细胞组、CD4⁺ T细胞组(P < 0.05,P < 0.01)。胚胎骨髓来源间充质干细胞与CD4⁺ T细胞共培养组Th17细胞数量明显高于CD4⁺ T细胞组(P < 0.01),但胚胎骨髓来源间充质干细胞本身并不表达白细胞介素17。表明胚胎骨髓来源间充质干细胞可促进人Th17细胞增殖。     

Mimotopes for tumor-specific T lymphocytes in human cancer determined with combinatorial peptide libraries

Mimotopes of a tumor-associated T cell epitope were determined using randomized and combinatorial peptide libraries and a CD8(+) T cell clone specific for the cutaneous T cell lymphoma cell line MyLa. Antigen recognition by this clone was found to be HLA-B8 restricted. More than 80 % of HLA-matched patients with cutaneous T cell lymphoma had mimotope-specific CD8(+) T cells in their peripheral blood. Mimotope-specific T cells isolated and expanded from a patient lysed MyLa cells in in vitro assays thus demonstrating their cytolytic capacity and tumor specificity. Mimotope vaccination of a patient without detectable mimotope-specific T cells induced frequencies of these cells of up to 1.82 % of the peripheral blood CD8(+) T cells.