Enhanced surface TCR replenishment mediated by CD28 leads to greater TCR engagement during primary stimulation

When T cells are stimulated with high concentrations of strong TCR agonist, engaged TCR are internalized and degraded, resulting in greatly reduced surface TCR levels for up to several days post-stimulation. It has been noted that surface TCR levels rise subsequently, even in the presence of continuing stimulation, but the role of CD28 co-stimulation in surface TCR replenishment has not been investigated. Here, we have examined the return of surface TCR following activation, the availability of these TCR for antigenic engagement and the role of CD28 in that process. We report that within 24 h of stimulation, the level of surface TCR expression becomes dependent on the degree of CD28 signaling provided during T cell activation. In addition, when cells are removed from stimulus after 24 h, surface TCR expression recovers to a stable level which exceeds that of unstimulated cells and is proportional to the degree of CD28 co-stimulation. TCR that replenish the plasma membrane during T cell activation can be down-regulated by receptor occupancy with the same efficiency as TCR on freshly stimulated cells. Thus, as a result of enhanced surface TCR replenishment, CD28-co-stimulated cells can engage and down-regulate more TCR than co-stimulation-deprived cells in the face of ongoing stimulation. Furthermore, engagement of newly expressed TCR on activated T cells re-induces CD69, suggesting participation of these replenishing TCR in continued T cell signaling. These data identify the augmentation of surface TCR replenishment during activation as a novel mechanism that likely contributes to the enhanced antigenic sensitivity of CD28-co-stimulated T cells.     

Role of CD28 co-stimulation in generation and maintenance of virus-specific T cells

Efficient induction of T cell responses is normally assumed to require both TCR-mediated signaling and engagement of co-stimulatory molecules, in particular CD28. However, the importance of CD28 co-stimulation in induction and maintenance of antiviral T cell responses is not clearly established. For this reason antiviral CD4(+) and CD8(+) T cell responses in CD28-deficient mice were studied using two different viruses [vesicular stomatitis virus and lymphocytic choriomeningitis virus (LCMV)]. Intracellular cytokine staining and/or MHC-peptide tetramers were used to enumerate antigen-specific T cells. In addition, we used DNA constructs encoding viral epitopes to probe the importance of the epitope itself. Our results reveal that while the context of antigen presentation (live virus versus DNA construct) is a critical factor in determining the requirement for CD28 co-stimulation, epitope and virus dose play little if any role. Direct visualization of antigen-specific cells also confirms the notion that CD28 is more critical for the generation of antiviral T(h)1 cells than for T(c)1 cells generated in response to the same virus (LCMV). Most importantly, the present study reveals that CD28 generally is essential for the host to respond optimally over a broad set of conditions, and our results may imply that the relatively CD28 independent activation of LCMV-specific CD8(+) T cells may represent an extreme situation related to the non-cytolytic nature of this virus allowing the delivery of a uniquely strong and prolonged signal 1.     

Comparison of thymocyte development and cytokine production in CD7-deficient, CD28-deficient and CD7/CD28 double-deficient mice

CD7 and CD28 are Ig superfamily molecules expressed on thymocytes and mature T cells that share common signaling 0mechanisms and are co-mitogens for T cell activation. CD7-deficient mice are resistant to lipopolysaccharide (LPS)-induced shock syndrome, and have diminished in vivo LPS-triggered IFN-gamma and tumor necrosis factor (TNF)-alpha production. CD28-deficient mice have decreased serum Ig levels, defective IgG isotype switching, decreased T cell IL-2 production and are resistant to Staphylococcus aureus enterotoxin-induced shock. To determine synergistic roles CD7 and CD28 might play in thymocyte development and function, we have generated and characterized CD7/CD28 double-deficient mice. CD7/CD28-deficient mice were healthy, reproduced normally, had normal numbers of thymocyte subsets and had normal thymus histology. Anti-CD3 mAb induced similar levels of apoptosis in CD7-deficient, CD28-deficient and CD7/CD28 double-deficient thymocytes as in control C57BL/6 mice (P = NS). Similarly, thymocyte viability, apoptosis and necrosis following ionomycin or dexamethasone treatment were the same in control, CD7-deficient, CD28-deficient and CD7/CD28-deficient mice. CD28-deficient and CD7/CD28-deficient thymocytes had decreased [3H]thymidine incorporation responses to concanavalin A (Con A) stimulation compared to control mice (P < or = 0.01 and P < or = 0.05 respectively). CD7/CD28 double-deficient mice had significantly reduced numbers of B7-1/B7-2 double-positive cells compared to freshly isolated wild-type, CD7-deficient and CD28-deficient thymocytes. Con A-stimulated CD4/CD8 double-negative (DN) thymocytes from CD7/CD28 double-deficient mice expressed significantly lower levels of CD25 when compared to CD4/CD8 DN thymocytes from wild-type, CD7-deficient and CD28-deficient mice (P < 0.05). Anti-CD3-triggered CD7/CD28-deficient thymocytes also had decreased IFN-gamma and TNF-alpha production compared to C57BL/6 control, CD7-deficient and CD28-deficient mice (P < or = 0.05). Thus, CD7 and CD28 deficiencies combined to produce abnormalities in the absolute number of B7-1/B7-2-expressing cells in the thymus, thymocyte IL-2 receptor expression and CD3-triggered cytokine production.     

Involvement of CD70 and CD80 intracytoplasmic domains in the co-stimulatory signal required to provide an antitumor immune response

CD70 and CD80 are co-stimulatory molecules which belong to the tumor necrosis factor family and the B7 family respectively. When they are co-expressed by gene-modified TS/A tumor cells, they provide an efficient protective and long-lasting T-dependent antitumor response. We first showed that when CD70 and CD80 were delivered in the tumor environment by gene-modified fibroblasts, but were not expressed by the tumor cells themselves, no antitumor response was observed. We next assessed whether the intracytoplasmic domains of CD70 and CD80 contribute to enhance the co-stimulatory activity necessary to induce effective T cell-tumor cell interactions and T cell-dependent antitumor response. TS/A cells were gene-modified to express different combinations of deleted (CD70Delta and CD80Delta) or full-length CD70 and CD80 co-stimulatory molecules. In vitro, the CD80 intracytoplasmic domain was required to regulate CD80 membrane redistribution by interacting with the actin cytoskeleton. The loss of the CD70 intracytoplasmic domain did not alter its ability to relocate on the surface membrane, but failed to co-stimulate T cell proliferation. In vivo experiments in syngeneic BALB/c mice showed that the CD70/CD80-TS/A and the CD70Delta/CD80-TS/A tumors were rejected via CD8 T cells, whereas CD70/CD80Delta-TS/A and CD70Delta/CD80Delta-TS/A tumors were not. The mice that rejected CD70Delta/CD80-TS/A tumors showed decreased protection against injection of parental TS/A cells when compared to mice which rejected CD70/CD80-TS/A tumors. These results showed that the intracytoplasmic domain of CD80 was critical for the effector phase of CD8 T cell-dependent tumor rejection and that the CD70 intracytoplasmic domain could mediate proliferative or surviving signals required for optimal effector/memory CD8 T cell generation.     

Gene-targeted B-deficient mice reveal a critical role for B cells in the CD4 T cell response

The role of B cells in promoting T cell responses is still controversial.In this study, we use JHD mice which have a targeted mutationin the J^H gene and are thus rendered deficient in B cells toaddress this issue. We show here that immunization of JHD micewith soluble antigen fails to prime CD4 T cells, for eitherclonal expansion or delivery of immunological help for antibodyresponses. This lack of CD4 T cell priming in JHD mice correspondsto a 3-to 9-fold lower co-stimulatory activity of antigen-presentingcells (APC) from the JHD mice, as measured by anti-CD3-inducedproliferative responses of CD4 T cells. This in turn is dueto a defect of APC from JHD mice in response to T cell-mediatedinduction of co-stimulatory activity. As the development ofmacrophages and dendritic cells is unaffected in the JHD mice,our results demonstrate that B cells play a critical role inCD4 T cell priming, possibly by delivering a critical co-stimulatoryactivity for clonal expansion of CD4 T cells.     

人CD137单抗诱导不同状态T细胞增殖和凋亡的研究

为了研究一种新的T细胞共刺激分子—CD137对不同状态T细胞增殖和凋亡的双重调节作用。采用3 H TdR掺入法测定T细胞增殖 ,用流式细胞仪测定细胞凋亡。结果显示 :(1)CD137单抗可与T细胞表面的CD137抗原结合 ,明显增强PHA刺激T细胞增殖 ,使T细胞增殖指数较PHA单独作用高 2～ 3倍 ,但CD137单抗单独不能刺激T细胞增殖 ;(2 )对于慢性活化的T细胞 ,CD137单抗可协同PHA诱导T细胞凋亡 ,使T细胞凋亡率从PHA单独作用的 19 2 0 %增加到 36 31% ,CD137单抗单独并不能诱导慢性活化T细胞凋亡。CD137单抗一方面可协同PHA刺激静止状态T细胞的增殖 ,另一方面可协同PHA诱导慢性活化T细胞凋亡 ,对T细胞起双重调节作用。     

特发性血小板减少性紫癜患者外周血共刺激分子的表达及与血小板抗体关系的临床研究

本研究检测特发性血小板减少性紫癜（ITP）患者外周血共刺激分子CD80、CD86和CD137的表达及血清血小板抗体（PAIgG）含量并探讨二者相关性及与血小板数量等疾病表现的关系,以期阐明共刺激分子在特发性血小板减少性紫癜发病及病情判断中的作用。分别应用免疫荧光法和流式细胞术检测48例ITP患者及40名正常人外周血单个核细胞（PBMNC）表面共刺激分子CD80、CD86和CD137的表达;应用双抗体夹心酶联免疫吸附法（ELISA）检测血清PAIgG含量。结果表明：ITP患者CD80、CD86和CD137的表达水平分别为（4．92±2．02）％,（8．68±4．25）％,（5．32±2．67）％,PAIgG平均含量为210±3．02ng／10^7PA,均明显高于正常对照组（2．01±0．75）％,（4．56±2．06）％,（1．37±1．25）％和20±1．13ng／10^7 PA（p〈0．01）。共刺激分子表达水平与PAIgG含量呈正相关（r=0．302,P〈0．05）,与患者血小板数量呈负相关（r=-0．369,P〈0．05）。结论：共刺激分子CD80、CD86和CD137是参与ITP发病和免疫反应的重要共刺激分子,其过度表达与ITP发病及临床病情密切相关。纠正其异常表达、调节免疫状态可能是ITP的治疗策略之一,具有重要的临床研究意义。     

中医体质与免疫共刺激分子CD40、CD28基因多态性的关联性研究

目的 研究中医体质与免疫共刺激分子单核苷酸多态性之间的相关性,探究中医体质分类理论与机体免疫系统功能间联系.方法 利用标准化中医体质量表结合中医四诊综合辨识,对359例志愿者进行中医体质辨识,利用聚合酶链式反应限制片段长度多态性(PCR-RFLP)方法鉴定其免疫共刺激分子CD40、CD28基因主要标签单核苷酸多态性位点(tagSNP)基因型.结果 对359例志愿者进行中医体质分类,发现CD40基因rs1883832位点基因型和等位基因在平和质和阳虚质中的分布频率存在统计学差异(P ＜0.05);CD28基因单体型GGCCTTCATAA与ACCTTTCATGA在平和质、气虚质和阳虚质三种中医体质中分布频率存在统计学差异(P＜0.05).结论 中医体质论与免疫共刺激分子基因单核苷酸多态性存在关联性,免疫共刺激分子基因单核苷酸多态性所造成的免疫功能个体差异可能是不同中医体质个体存在疾病易感性差异的理论依据之一.     

PD-1/PD-L1在非小细胞肺癌中的研究进展及展望

PD-1(programmed cell death-1,程序性死亡受体1)与其配体PD-L1(programmed cell death-ligand 1,程序性死亡配体1)属于CD28/B7家族,是一对共刺激分子,具有负性调控作用。PD-1通过与其配体PD-L1结合调节肿瘤的微环境,使肿瘤细胞免于机体免疫系统的监视和清除。目前已有较多研究显示PD-1/PD-L1在非小细胞肺癌组织中的表达水平与患者的临床病理因素及预后存在显著的相关性。在非小细胞肺癌的治疗领域,以PD-1/PD-L1为代表的免疫治疗成为继手术治疗、化疗、放疗、分子靶向治疗之后的新焦点。PD-1/PD-L1抑制剂在一系列非小细胞肺癌临床试验中也显示出了巨大的临床潜力。本文就PD-1/PD-L1的生物学结构及其在非小细胞肺癌中的作用机制、研究进展及展望作一综述。     

PDL1修饰树突状细胞对大鼠同种异体肾移植存活的影响

目的观察激活负性共刺激信号PD1-PDL1对延长移植肾存活的效能。方法以PDL1Ig基因重组腺病毒为载体,将PDL1Ig基因转入BN大鼠树突状细胞(DC)细胞同时输注受体,以Lewis大鼠为受体,行同种肾移植为转染组,并以未转染DC输注为对照组;观察移植肾存活时间和术后肾功能变化。结果转染组移植肾存活(41±7.6)d,较对照组移植肾存活时间(8.6±1.2)d明显延长;移植组术后血清肌酐较同期对照组明显为低。结论PDL1基因修饰的DC可以明显延长移植肾存活时间。