闭合性创伤后T细胞膜流动性的改变及其与T细胞功能的关系

本研究观察了闭合性创伤小鼠Ｔ细胞膜流动性的变化及其与Ｔ细胞功能之间的关系。结果显示，创伤后Ｔ细胞质膜、线粒体膜、微粒体膜流动性均降低，且这一变化同创后Ｔ淋巴细胞转化降低、白细胞介素２（ＩＬ－２）产生减少、ＩＬ－２受体表达受抑、ＩＬ－２介导的淋巴细胞增殖反应低下显著相关。     

索拉非尼抑制T淋巴细胞的分子机制研究

目的 阐明索拉非尼抑制正常人外周血T淋巴细胞的分子机制.方法 羧基荧光素二醋酸盐琥珀酰亚胺酯(5,6-carboxyfluorescein diacetate succinimidyl ester,CFSE)增殖试验和MTS[3-(4,5-二甲基噻唑-2-基)-5-(3-羧甲酯基)-2-(4-磺苯基)-2H-四唑(金翁),内盐]法检测索拉非尼对T细胞的增殖和生存能力的影响;Annexin V和PI双染法检测T细胞的凋亡;FACS检测细胞周期和CD25、CD69的表达,Western blot检测细胞周期蛋白的表达;ELISA法检测IL-2的水平;采用苦基氯(picryl chloride,PCL)诱导的迟发性过敏反应(DTH)模型,检测索拉非尼对小鼠体内T细胞免疫应答的影响.结果 索拉非尼剂量依赖性的抑制T淋巴细胞的增殖,却不影响T细胞的凋亡,它可以使T细胞周期阻滞在G_0/G_1期.索拉非尼可以抑制T细胞CD25、CD69的表达,抑制IL-2的分泌,在体内索拉非尼能够抑制PCL诱导DTH模型.结论 索拉非尼可以抑制外周血T淋巴细胞的增殖和活化,临床上长期用药时可能会导致机体的免疫抑制作用.     

恶性骨肿瘤患者巨噬细胞功能、NK细胞活性和T细胞亚群的变化

我们检测了３４例恶性骨肿瘤患者的巨噬细胞功能、ＮＫ细胞活性和Ｔ细胞亚群的变化,可为临床上恶性骨肿瘤的免疫治疗及病情判定,提供一定的实验依据。１材料和方法１．１研究对象患恶性骨肿瘤患者３４例,均为经ＥＣＴ检查而确诊的住本院患者,包括：骨肉瘤２０例、滑膜肉瘤１例、软骨肉瘤２例、恶性纤维组织细胞瘤１例、转移癌３例及骨巨细胞瘤７例。其中男２３例,女１１例,年龄１０～８５岁平均３０．０岁。此外,另选择本单位健康人３２例均经查体证实,作为对照组,平均年龄３５．５岁。１．２方法①Ｔ细胞亚群的检测〔１〕：取患者…     

T淋巴细胞归巢分子机制的研究进展

淋巴细胞归巢（LH）是指血液循环中淋巴细胞选择性穿越毛细血管后高内皮微静脉定向迁移并进入外周免疫器官或特定组织区域.淋巴细胞与各组织、器官血管内皮黏附分子的相互作用是淋巴细胞归巢的分子基础.淋巴细胞的归巢过程是一个多种黏附分子参与并受各种因素调节的复杂过程.因而T淋巴细胞归巢分子机制很重要.     

CD28以外的T细胞协同刺激分子及其功能

CD2 8是协同 TCR诱导 T细胞产生高水平 IL - 2 ,促进 T细胞生存及防止 T细胞凋亡的主要协同刺激分子 ,但并非所有 T细胞均表达 CD2 8,提示 T细胞活化尚有其它协同刺激途径的存在。最近研究表明 ,其它一些协同刺激分子 ,如 4 - 1BB、OX4 0、ICOS、L FA- 1等在不同的 T细胞亚群和 T细胞活化的不同阶段 ,呈现不同的表达和介导不同的生物学功能     

体外抗原诱导小鼠T细胞对NK细胞功能的抑制及其可能分子机制研究

目的:为了探讨抗原诱导T细胞抑制对NK细胞功能的抑制及其可能机制.方法:在体外,分别利用抗原、抗原 抗CD80抗体、刀豆蛋白A、抗CD3抗体先行诱导小鼠T细胞,并于诱导后24、48、72、96小时分别引入NK细胞,通过51Cr释放试验动态观察NK细胞的功能状况,激光共聚焦显微镜观察T细胞抑制NK细胞功能的作用模式,同时用RT-PCR方法检测经诱导的T细胞Bc1与Bc2表达情况.结果:①各组在24、48、72小时相点,上清液对NK功能均无显著影响,而到96小时,抗原激活组与抗原 抗CD80抗体诱导组两组上清液对NK细胞的杀伤能力表现出促进作用,与其它各时相点相比有统计学意义(P＜0.05),且后者低于前者,二者相比也具统计学意义(P＜0.05).②刀豆蛋白A与抗CD3抗体刺激组T细胞从T细胞接受刺激24小时起至96小时,均表现了抑制NK细胞功能.抗原 抗CD80抗体耐受诱导组与抗原激活组T细胞则是在诱导后48小时方表现出对NK细胞的抑制,与其它两组相比差异具有统计学意义(P＜0.05).③只有同种抗原刺激48小时的细胞培养物可检出Bc1、Bc2.其他时相、其他细胞刺激剂作用下均未能检出Bc1,Bc2的表达.结论:经诱导的T细胞能以直接接触的方式抑制NK细胞功能,但不同诱导剂活化的T细胞引起NK细胞抑制的机制可能不同.小鼠HLA-G的类似物-Bc1与Bc2的表达与抗原信号有关,抗原诱导的小鼠T细胞可能通过表达Bc1与Bc2来抑制NK细胞的功能,从而参与移植耐受的形成,这一结论提示这类非经典MHC Ⅰ类分子有可能成为移植耐受的检测标志.     

输精管结扎6月与25月家兔T淋巴细胞与巨噬细胞功能的观察

本文应用输精管结扎６月，２５月家兔的动物模型，观察了海马培养上清胸腺细胞调节因子活性，胸腺细胞自身增殖反应性与脾脏的Ｔ，Ｂ细胞功能，及外周血巨噬细胞功能的变化。结果表明，（１）海马培养上的清胸腺细胞调节因子活性，胸腺细胞自身增殖反应，输精管结扎６月组和输精管结扎２５月组与相应的对照组比较差异均无显著性；（２）ＣｏｎＡ诱导的脾细胞增殖反应性和ＩＬ－２，ＩＬ－６活性，输精管结扎６月组均呈低值，且ＩＬ－     

创伤后大鼠pMΦ分化表型及吞噬功能的变化

目的 观察创伤后不同时相大鼠腹腔巨噬细胞分化表型与吞噬功能的动态变化。方法 采用创伤失血模型，运用流式细胞仪分析大鼠巨噬细胞标志性抗原ED1、分化抗原ED2的表达情况，并用无色孔雀绿法测定大鼠腹腔巨噬细胞吞噬功能变化。结果 创伤后大鼠pM中分化表型ED2表达比例降低，于创伤后12h至最低，伤后第2天有短暂升高，但严重创伤后至第7天仍未完全恢复至正常，无单纯的ED2表达；无色孔雀绿法测定创伤后非特异吞噬功能下降，于创伤后第1天达最低点，LPS刺激可持续减弱吞噬功能。结论 创伤影响巨噬细胞的分化状态，伴随表型改变而出现的分化状态不同可能是吞噬功能减低的原因之一。     

Agonist-induced T cell receptor down-regulation: molecular requirements and dissociation from T cell activation

T cell receptor (TCR) down-regulation is a consequence of specific receptor engagement and plays an important role in modulating the T cell response. We have investigated the role of protein kinase C (PKC) and protein tyrosine kinases (PTK) in the induction of TCR down-regulation. We report that the mutation of S126 in the CD3-γ chain that is known to inhibit phorbol-12-myristate 13-acetate-induced TCR down-regulation does not affect down-regulation induced by a specific agonist. In addition, agonist-induced TCR down-regulation is not affected by blockade or depletion of PKC, neither by blockade or lack of PTK, while the same treatments efficiently interfere with T cell activation. These results demonstrate that TCR down-regulation is induced by early events which follow specific engagement by an agonist and can be dissociated from those required for full T cell activation.     

The effect of rapamycin on T cell development in mice

Rapamycin (RAPA) is a strong immunosuppressant with a chemical structure similar to that of FK506, although it acts by a mechanism different from both FK506 and cyclosporin A. The effect of RAPA on T cell development in mice was investigated in this study. RAPA caused significant thymic atrophy. The major histological change in the RAPA-treated thymus was thinning of the cortex. No other apparent damage in the cortex or medulla was observed. Consistent with these histological findings, in vivo thymocyte cycling was blocked by RAPA before the S phase, and the ex vivo and in vitro proliferation of the thymocytes was also strongly repressed by the drug. According to electron microscopy and DNA fragmentation assays, RAPA did not induce apoptosis. These results indicate that the repressed thymocyte proliferation is a major mechanism causing RAPA-induced thymic atrophy. Further, RAPA had no effect on thymocyte apoptosis induced by anti-CD3 or ionomycin, and the drug did not interfere with deletion of CD4⁺8⁺ thymocytes or peripheral Vβ6⁺ T cells induced by anti-CD3 or Mls-1^a, respectively. These data suggest that RAPA does not hamper the negative selection. There was a relative increase in the CD3^? fraction of the de novo developing CD4 and CD8 double-positive (DP) thymocytes in the RAPA-treated mice. Moreover, there were relative increases of the CD3^? fractions of the CD4 or CD8 single-positive (SP) cells in both the thymi and periphery. The generation of the CD3^? SP under the influence of RAPA could be used as a useful model for further study of the function and signal transduction of these CD3-defective SP cells.     

血必净对活化诱导T细胞凋亡的调节

目的 观察活化诱导对脾脏T淋巴细胞凋亡、凋亡相关基因mRNA表达及caspase3活性的影响,以及活血化瘀中药的调节作用.方法 提取BALB/c小鼠脾脏T淋巴细胞并培养,以Con A+IL-2诱导T细胞活化凋亡,MTT法检测细胞增殖活性,流式细胞仪检测细胞凋亡率,RT-PCR检测Fas、FasL、Bcl-2、Bax、IL-2 mRNA表达水平,分光光度法测定caspase3酶活性,并观察活血化瘀中药对上述各项指标的影响.结果 活化T淋巴细胞于诱导18h后凋亡率明显增加,于诱导6h时未见FasL、Bax mRNA表达,Fas、Bcl-2 mRNA表达无明显变化;于诱导18 h后Fas、FasL、Bax mRNA表达升高,Bel-2 mRNA表达下降,caspase3活性增高.活血化瘀中药可降低T细胞凋亡,并可分别降低Fas、FasL、Bax mRNA表达,提高Bcl-2 mRNA表达,减轻easpase3酶活性.在活化诱导早期(6 h)促进T淋巴细胞内IL-2 mRNA表达,在晚期(18 h)减少IL-2 mRNA表达.结论 过度活化是脾脏T淋巴细胞异常凋亡的诱发因素,而凋亡的发生与Fas、FasL、Bax、Bcl-2 mRNA表达的改变有关.活血化瘀中药可通过调节IL-2及凋亡相关基因mRNA表达而减轻脾脏T淋巴细胞凋亡,同时可以促进T淋巴细胞的增殖活性.     

Fasudil mediates cell therapy of EAE by immunomodulating encephalomyelitic T cells and macrophages

Although Fasudil has shown therapeutic potential in EAE mice, the mechanism of action are still not fully understood. Here, we examined the immunomodulatory effect of Fasudil on encephalitogenic mononuclear cells (MNCs), and tested the therapeutic potential of Fasudil‐treated MNCs in active EAE. Fasudil inhibited expression of CCL20 on T cells and migration of T cells, decreased CD4⁺IFN‐γ⁺ and CD4⁺IL‐17⁺ T cells, but increased CD4⁺IL‐10⁺ and CD4⁺TGF‐β⁺ T cells. Fasudil reduced expression of CD16/32 and IL‐12, while elevating expression of CD206, CD23, and IL‐10. Fasudil also decreased levels of iNOS/NO, enhanced levels of Arg‐1, and inhibited the TLR‐4/NF‐κB signaling and TNF‐α, shifting M1 macrophage to M2 phenotype. These modulatory effects of Fasudil on T cells and macrophages were not altered by adding autoantigen MOG_35–55 to the culture, i.e., autoantigen‐independent. Further, we observed that, in vitro, Fasudil inhibited the capacity of encephalitogenic MNCs to adoptively transfer EAE and reduced TLR‐4/p‐NF‐κB/p65 and inflammatory cytokines in spinal cords. Importantly, Fasudil‐treated encephalitogenic MNCs exhibited therapeutic potential when injected into actively induced EAE mice. Together, our results not only provide evidence that Fasudil mediates the polarization of macrophages and the regulation of T cells, but also reveal a novel strategy for cell therapy in MS.     

B and T cell tolerance and autoimmunity in autoantibody transgenic mice

One hallmark of systemic lupus erythematosus (SLE) is the presence of autoantibodies directed at a diverse group of proteins of the U1/Sm small nuclear ribonucleoprotein particles (snRNP). Patients with SLE and murine models of this disease generate high titers of affinity mature, isotype-switched autoantibodies characteristic of T cell-dependent immune responses. In this investigation, we made use of anti-snRNP Ig transgenic mice (2-12 Tg) to track regulation of autoreactive B cells in normal and autoimmune-prone mice. Autoantibody studies demonstrated that the regulation of anti-snRNP B cells is intact in non-autoimmune Tg mice, but not in MRL-lpr/lpr mice. We further utilize autoreactive Tg B cells as antigen-presenting cells (APC) and individual snRNP peptides to assess the presence of autoreactive T cells in the repertoire of non-autoimmune and MRL-lpr/lpr mice. We found that Tg B cells can direct specific T cell tolerance in a non-autoimmune-prone (C57Bl/6) background, whereas the same autoantibody transgene in MRL-lpr/lpr mice drives T cell autoimmunity. Moreover, Tg B cell APC could stimulate autoreactive T cells from wild-type (non-Tg) C57Bl/6 mice, indicating a lack of tolerance induction in the absence of the autoantigenic-presenting B cells. Thus, we have defined dual roles for autoantigen-presenting B lymphocytes in stimulating self-reactive T cells that inhabit the normal repertoire or, under some conditions, providing tolerance signals.     

Suppression of autologous peripheral blood mononuclear cell proliferation by alveolar macrophages from young infants

Maintenance of lung homeostasis involves a complex interaction between T lymphocytes and alveolar macrophages (AM), in which AM suppress pulmonary T cell proliferation to antigenic stimuli. To assess whether AM-mediated suppression is attenuated in healthy young infants, AM and peripheral blood mononuclear cells (PBMC) were sampled prior to elective surgery. Children were divided into <4 months of age (Group I) and >4 months (Group II). Autologous PBMC and AM were co-cultured in vitro with phytohaemaglutinin (PHA) at AM : PBMC ratios ranging from 2:1 to 1 : 5. Methyl-tritiated thymidine was added after 48 h and uptake determined at 72 h. Percentage suppression or enhancement of PBMC proliferation by AM was determined relative to proliferation of PBMC with PHA. To determine the role of soluble factors of suppression, cell-free supernatants from paediatric AM and PBMC co-cultures were added to PHA-stimulated adult PBMC. The median age was 3 months for Group I (n = 9) and 7 years 2 months (n = 13) for Group II. Percentage suppression of PBMC proliferation was attenuated in Group I (versus Group II) at AM : PBMC ratios of 2:1 (median 78 versus 92, P< 0 x 05) and 1 : 1 (45 versus 87, P< 0 x 01). Cell-free supernatants from Groups I and II suppressed proliferation of adult PBMC, but there was no difference in suppression between the age groups. We conclude that suppression of autologous PHA-stimulated PBMC proliferation by AM is attenuated in young infants, and this immaturity is not explained by reduced release of soluble factors.     

Leishmania-infected macrophages sequester endogenously synthesized parasite antigens from presentation to CD4+ T cells

CD4⁺ T cell lines raised against the protective leishmanial antigens GP46 and P8 were used to study the presentation of endogenously synthesized Leishmania antigens by infected cells. Using two different sources of macrophages, the 14.07 macrophage cell line (H-2^k) which constitutively expresses major histocompatibility complex (MHC) class II molecules, and elicited peritoneal exudate cells, we found that cells infected with Leishmania amastigotes presented little, if any endogenously synthesized parasite antigens to CD4⁺ T cells. In contrast, promastigote-infected macrophages did present endogenous parasite molecules to CD4⁺ T cells, although only for a limited time, with maximal presentation occurring within 24 h of infection and decreasing to minimal antigen presentation at 72 h post-infection. These observations suggest that once within the macrophage, Leishmania amastigote antigens are sequestered from the MHC class II pathway of antigen presentation. This allows live parasites to persist in infected hosts by evading the activation of CD4⁺ T cells, a major and critical anti-leishmanial component of the host immune system. Studies with drugs that modify fusion patterns of phagosomes suggest that the mechanism of this antigen sequestration includes targeted fusion of the parasitophorous vacuole with certain endocytic compartments.     

CD28 can promote T cell survival through a phosphatidylinositol 3-kinase-independent mechanism

Phosphatidylinositol 3(Pl3)-kinase is implicated in various biological responses, including protection from apoptosis, although its role in antigen-induced T cell death and the molecular effectors it triggers remains ill-defined. Here, we investigated the role of Pl3-kinase activity in the prevention of T cell receptor/CD3-induced cell death by CD28. Pl3-kinase inhibitors blocked the up-regulation of Bcl-X_L by CD28, without impairing the prevention of T cell receptor/CD3-triggered apoptosis by CD28, hence showing the existence of a cell-survival pathway independent of Pl3-kinase activity and up-regulation of Bcl-X_L. Instead, we show that up-regulation of FasL which is instrumental in CD3-induced apoptosis was prevented upon CD28 co-stimulation. These results indicate that Pl3-kinase couples CD28 to Bcl-X_L up-regulation and provide a molecular basis for the role of CD28 in cell survival through a Pl3-kinase-independent mechanism including FasL down-regulation.     

年龄、性别在幼鼠胸腺T细胞增殖中的作用

目的：分析年幼小鼠性别、年龄与CD3、CD4、CD8免疫参数表达的关系。方法：分离3—9周小鼠胸腺、脾脏T淋巴细胞，FACS分析细胞表面CD3、CD4、CD8的表达；CFSE标记后细胞，加入ConA、抗TCR抗体、PMA IONO刺激剂培养72小时，流式细胞仪分析的方法比较不同刺激剂作用下雌、雄小鼠T细胞的增殖。结果：结果胸腺和脾脏表面CD3、CD4、CD8的表达与性别无明显关系，6-9周小鼠脾脏CD3^ 细胞明显多于3—4周小鼠；胸腺细胞的增殖与年龄、性别无关，而3周雌性小鼠脾细胞对PMA IONO刺激后的增殖应答比雄性明显，4-6周雄性小鼠脾细胞的增殖能力强于雌性小鼠，7—8周雌性小鼠脾细胞对抗TCR抗体的应答能力明显减小。结论：免疫系统的雌雄异型可能早于青春期。     

The degree of CD8 dependence of cytolytic T cell precursors is determined by the nature of the T cell receptor (TCR) and influences negative selection in TCR-transgenic mice

Although much has been learned about CD8 structure-function properties, it has so far not been tested whether the nature of the TCR is sufficient to transfer the property of CD8 dependence versus non-dependence to CD8⁺ cytotoxic T lymphocytes (CTL) and their precursors differentiating in T cell receptor (TCR)-transgenic (Tg) mice. In the present study, we compared the characteristics of dependence on CD8 for stimulation of CTL precursors and antigen-specific cytolysis by CD8⁺ T cells from two TCR-Tg mice expressing respectively the TCR (Tg) from a “CD8-dependent” and from a “CD8-independent” CTL clone, which were both reactive against the H-2K^b alloantigen and originated from H-2^k mice. The results indicate that the property of the Tg⁺CD8⁺ cells from H-2^k TCR-Tg mice corresponds to that of the CTL clone of origin, demonstrating that it is linked to the nature of the TCR. Consistent with this property, Tg⁺CD4⁺ cells could also differentiate into H-2K^b-specific CTL when originating from the “CD8-independent”, but not from the “CD8-dependent” Tg-TCR. The influence of the property of “CD8 dependence” on negative selection occurring in TCR-Tg H-2^klb mice was apparent at two levels: (i) in the thymus, the extent of deletion was much more pronounced for the “CD8-independent” TCR-Tg mice; (ii) in the periphery, Tg^+(hi) cells with low to negative CD8 expression were present for the “CD8-dependent” Tg-TCR, whereas only Tg⁺CD4^?CD8^? cells with low surface Tg-TCR and CD3 expression were found for the “CD8-independent” Tg-TCR, indicating that Tg⁺CD4^?CD8^? cells are susceptible to tolerance induction involving TCR/CD3 surface down-modulation. Furthermore, different in vitro conditions led to H-2K^b-induced stimulation of Tg⁺CD4^?CD8^? cells to differentiate into CTL detected in an anti-TCR clonotypic monoclonal antibody redirected cytolysis assay. Culture in interleukin-2 of H-2^klb Tg⁺CD4^?CD8^? cells was sufficient to induce CTL activity in the “CD8-independent” model, whereas stimulation with cells which overexpressed H-2K^b was required in addition to interleukin-2 to induce CTL differentiation in the “CD8-dependent” model. These data suggest that peripheral Tg⁺CD4^?CD8^? cells present in a situation of in vivo tolerance to H-2K^b can still be triggered by H-2K^b with a sensitivity correlated with the degree of CD8 dependence.