IL-2和IL-15协同调节T细胞的增殖和LAK细胞的杀伤活性

目的 探讨IL-2和IL-15在免疫调控和应答中的协同作用。方法 IL-2、IL-15和IL-2 IL-l5组刺激CTLL细胞的增殖，^3H—TdR掺入法测cpm值；IL-2、IL-15和IL-2 IL-l5组诱导PBMC中NK和LAK细胞的发育，4h^51Cr释放实验检测对K562和LiBr细胞的杀伤活性。结果 IL-2和IL-15都能诱导CTLL认细胞的增殖和NK、LAK细胞的杀伤活性，IL-2和IL-15可明显协同上调CTLL认的增殖和LAK细胞的杀伤活性。结论 IL-2和IL-15在免疫调控和应答中有一定的协同作用，为细胞因子联合应用于临床治疗肿瘤等疾病提供了实验依据。     

吸入糖皮质激素对哮喘儿童外周血Foxp3~+调节性T细胞及IL-2、IL-6的影响及其机制研究

研究吸入丙酸氟替卡松(FP)对哮喘患儿外周血单个核细胞(PBMC)中CD4+Foxp3+调节T细胞、细胞因子IL-2、IL-6以及转录因子STAT5的影响。以30例确诊为支气管哮喘的患儿为研究对象,随机分为未治疗哮喘组(15例)、吸入FP缓解组(15例),10例同期正常儿童为对照组。流式细胞仪检测外周血PBMC中的CD4+Foxp3+调节T细胞比率,ELISA检测血浆或培养上清中IL-2、IL-6细胞因子水平,Western blot检测PBMC中磷酸化及非磷酸化STAT5的水平。结果1.未治疗哮喘组PBMC中CD4+Foxp3+T细胞百分率在PHA刺激培养前后均明显低于正常对照组,吸入FP缓解组明显升高,与正常对照组没有差异;各组刺激后CD4+Foxp3+T细胞百分率均有升高,吸入FP缓解组、正常对照组分别升高约1.89、2.01倍,而未治疗哮喘组升高仅1.56倍;2.未治疗哮喘组血浆中IL-6水平明显高于正常组及吸入FP缓解组,而IL-2水平没有明显差异;3.PHA刺激24 h后未治疗哮喘组磷酸化STAT5(p-STAT)表达水平明显低于吸入FP组及正常对照组,而各组STAT5表达水平没有明显差异,结论吸入FP能增加哮喘患儿外周血PBMC中CD4+Foxp3+调节T细胞数量,其机制可能与降低血浆IL-6,上调STAT5磷酸化水平有关。     

sIL—2R释放与T细胞活化的相关性研究

本实验利用双抗体夹心ＥＬＩＳＡ、流式细胞测量术及淋巴细胞增殖试验观察了可溶性ＩＬ－２受体（ｓＩＬ－２Ｒ）释放与细胞膜ＩＬ－２受体（ｍＩＬ－２Ｒ）表达及淋巴细胞增殖反应的相关性。结果显示：①在不更换培养液的情况下，ＰＨＡ刺激的淋巴细胞培养上清中的ｓＩＬ－２Ｒ释放与ｍＩＬ－２Ｒ表达在７２ｈ内呈正相关（ｒ＝０．９４，Ｐ〈０．０５），之后，ｍＩＬ－２Ｒ的表达逐渐减少，而ｓＩＬ－２Ｒ水平则继续缓慢增加，呈负     

IL－12对肿瘤病人PBMC增殖和杀瘤活性的实验研究

主要观察了ＩＬ－１２与ＩＬ－２联合对健康人和肿瘤病人ＰＢＭＣ增殖和体外杀瘤活性的影响。实验结果表明，单独应用ＩＬ－１０２对肿瘤病人ＰＨＡ活化ＰＢＭＣ增殖活性很小，如与低剂量ＩＬ－２合用，则可明显促进其增殖效应，表明ＩＬ－２可增强ＩＬ－１２的作用。与单用ＩＬ－１２相比，当ＩＬ－１２与低浓度ＩＬ－２合用时，健康人和肿瘤病人ＰＢＭＣ对Ｋ５６２细胞和Ｒａｊｉ细胞的杀伤活性均明显增强，且对Ｋ５６２细胞株杀伤活性明显强于对Ｒａｊｉ细胞株的杀伤活性。上述结果表明，ＩＬ－１２与ＩＬ－２合用对健康人和肿瘤病人ＰＢＭＣ增殖及杀瘤活性影响的格局基本一致。本实验为进一步研究ＩＬ－１２协同其它因子增强肿瘤病人ＬＭ和ＴＩＬ等免疫细胞抗肿瘤效应并进而使ＩＬ－１２过渡到临床肿瘤病人治疗提供了重要的实验依据。     

CD25含量与T辅助细胞对IL-2的促增殖反应性

目的 研究不同激活状态下T辅助细胞(Th)中IL-2R各亚基的表达变化及对IL-2反应的相关性.方法 以包被的抗CD3抗体激活Th1细胞,同时设立未激活对照.3H掺入法测定其对IL-2的促增殖反应;real-time PCR检测IL-2R各亚基编码基因的表达;流式细胞术检测CD25及CD122的含量;以I125标记的IL-2检测不同状态的Th1细胞对IL-2的亲和力.脂质体法转染IL-2Rα siRNA至激活的Th1细胞,比较不同CD25含量时Th1细胞对IL-2的反应性.自小鼠体内分离CD4细胞,以抗CD3抗体激活后在不同时间点收集细胞,比较它们的CD25含量及对1L-2的反应性.结果 较未激活对照相比,激活的Th1细胞中IL-2Rα亚基,即CD25的表达显著增加,而其他两个亚基则无变化;同时伴有对IL-2亲和力的升高,但对IL-2的促增殖反应性却显著降低.以siRNA适当下调IL-2Rα基因的表达则可显著提高激活的Th1细胞对IL-2的反应性.虽然激活后的na(i)ve CD4细胞对IL-2的反应性明显增加,但却不与CD25的含量及激活度正相关.最高的IL-2反应性出现在低度激活的CD4细胞中.结论 CD25在激活后的Th细胞中显著增多,并可通过自身数量的变化来调节所在细胞对IL-2的促增殖反应性.适度高表达的CD25可使靶细胞具有最高的IL-2反应性,过度表达的CD25则可导致对IL-2反应性的降低.     

人重组C3片段通过IL-2自分泌效应对CTLL-2细胞产生促增殖作用

目的研究补体Ｃ３片段的体外生物学活性及其作用机理，进一步探讨Ｃ３片段与免疫细胞的关联。方法利用重组ＤＮＡ技术表达纯化Ｃ３活性片段（命名为Ｃ３３），观察其对ＩＬ－２依赖性的小鼠杀伤性Ｔ细胞株ＣＴＬＬ－２细胞的促增殖效应，并通过抗体封闭途径和分子杂交技术研究Ｃ３３作用的机理。结果发现Ｃ３３蛋白对ＣＴＬＬ－２细胞具有明显的促增殖作用，并呈剂量依赖关系；这一作用能够部分地被抗小鼠ＣＤ１１ｂ抗体所封闭，能够完全被抗小鼠ＩＬ－２抗体所封闭；分子杂交显示Ｃ３３蛋白能够明显刺激ＣＴＬＬ－２细胞的ＩＬ－２ｍＲＮＡ表达。结论人重组Ｃ３片段Ｃ３３可通过ＩＬ－２的自分泌效应对ＣＴＬＬ细胞产生促增殖作用，补体受体ＣＲ３参与这一作用。     

老龄人群血浆IL-1活性和T细胞功能的随龄变化

以６２０例一汽集团离退休及在职男性职工为研究对象,检测外周血Ｔ淋巴细胞丝裂原反应性和血浆ＩＬ－２及ＩＬ－１活性,研究血浆ＩＬ－１活性和Ｔ细胞功能的随龄变化及二者相关关系。采静脉血５ｍＬ,肝素抗凝,无菌分离血浆,－３０℃冷冻保存,用于如下指标的检测。（...     

牛大力对抗体及IL-2产生的影响

<正>牛大力是一种具有补虎润肺、强筋活络功能的中药,临床已证明对多种慢性疾病有治疗作用.如对风湿性关节炎、肺结核、慢性支气管炎、慢性肝炎、白带等均有一定疗效.本文进一步研究了牛大力对实验小鼠B淋巴细胞分泌特异性抗体及T淋巴细胞产生白细胞介素2的免疫调节作用.1 材料和方法1.1 材料牛大力(Millettia speciosa champ)产于广东(由湛江吉民药业有限公司林振副研究员鉴定提供).~3H-TdR购自上海原子核研究所,CTLL-2细胞株(预防医学科学院病毒所),RPMI-1640培养液及ConA(Sigma公司产品).     

白细胞介素2与调节性T细胞介导的免疫耐受

调节性T细胞(Tregs)可以诱导机体对自身抗原和过敏原产生免疫耐受,从而维持机体免疫稳态.白细胞介素(IL)-2是一种具有双向免疫调节作用的糖蛋白,其不仅可以增强免疫反应,更重要的是可以维持Tregs的稳定及其介导的免疫耐受.研究IL-2与Tregs功能活性及其介导的免疫耐受的关系对更好地了解免疫相关性疾病的发病机制和指导临床正确使用IL-2或IL-2拮抗剂具有重要意义.     

Caspase在T细胞活化与增殖中的作用

半胱氨酸天冬氨酸特异性蛋白酶( caspases)是一类同源蛋白酶,主要参与细胞的凋亡过程.近年来的研究表明,该家族部分成员在T细胞活化与增殖中亦发挥重要作用,主要体现在以下3个方面:T细胞活化过程中检测到多种caspases活性;caspase-8失活后导致了人与鼠T细胞免疫的缺陷;体外实验中,caspase抑制剂能够阻断抗CD3单抗及超抗原引起的T细胞的活化与增殖.目前,部分体内实验已经证实了caspase抑制剂抗免疫反应及抗炎症反应的有效性,这为免疫性及炎症性疾病提供了新的治疗思路,为研制新型抗免疫、抗炎药物提供了依据.     

银杏内酯B对小鼠T淋巴细胞体外活化、增殖与凋亡的影响

目的银杏内酯B(GB)是已知的天然而强效的血小板激活因子(PAF)受体(PAFR)拮抗剂,本文研究GB对小鼠T淋巴细胞活化、增殖及凋亡3大体外行为的影响,初步探讨其潜在的免疫调节作用与机制,从而为临床应用提供可靠的实验依据。方法分离小鼠淋巴细胞,培养前以不同浓度的GB预孵;以刀豆蛋白A(Con A)诱导细胞的活化与增殖,以荧光抗体双色标记(anti-CD3 mAb-PE/anti-CD69 mAb-FITC、anti-CD3 mAb-PE/anti-CD25 mAb-FITC)结合流式细胞术分别检测T淋巴细胞早、中期活化标志--表面抗原CD69与CD25的表达,以活体荧光染料CFDA-SE标记结合流式细胞术检测细胞增殖并用MTT法佐证;以地塞米松(Dex)诱导细胞的凋亡,以荧光染料双色标记[DiOC6(3)/PI]结合流式细胞术区别凋亡、死亡与活细胞群。结果 Con A刺激后6 h和24 h,T细胞早期活化标志CD69和中期活化标志CD25分别大量表达,Con A刺激后48 h活化的T细胞增殖至第4代,而经终浓度为5、10、20μmol/L GB在Con A刺激前对细胞进行4 h预孵处理可以显著下调T细胞表面抗原CD69、CD25的表达并能有效抑制细胞增殖。Dex诱导后12 h细胞凋亡和死亡群显著增大,而经同样的GB预孵处理可以在一定程度上抑制细胞的凋亡进程,对细胞凋亡显示出一定的保护作用。结论 GB能有效抑制小鼠T淋巴细胞的活化与增殖,并且对细胞凋亡起到一定的保护作用,凭借GB对T细胞此3大行为出色的调节作用,理应将其作为天然的免疫抑制剂候选者进行更深入的研究。     

Histamine increases anti-CD3 induced IL-5 production of TH2-type T cells via histamine H2-receptors

Besides its proinflammatory functions histamine released from basophils and mast cells during immediate-type hypersensitivity reactions is known to inhibit several lymphocyte functions like IL-2 and -IFN production. Recently, it has been shown that T helper cells of type 2 phenotype (T_H2) represent the T cell fraction which may play a pivotal role in the promotion of the allergic inflammatory eosinophilic late-phase reaction by secretion of cytokines, especially IL-4 and IL-5. We have investigated the effect of histamine on anti-CD3 induced IL-4 and IL-5 production by T_H2 cells. Histamine in concentrations between 10^–7 and 10^–5 mol/l concentration-dependently increased anti-CD3 induced IL-5 production up to 120%, whereas IL-4 production was not affected. The activity of histamine in increasing IL-5 production was mimicked by the H₂-receptor agonist dimaprit. Histamine induced increase in IL-5 production was inhibited by histamine H₂-receptor antagonists, but remained unaffected by H₁- or H₃-receptor antagonists. Administration of forskolin which directly stimulates the production of cAMP, the second messenger of the H₂-receptor, also resulted in an increase in anti-CD3 induced IL-5 production. These results indicate that the histamine-mediated increase in anti-CD3 induced IL-5 production is mediated via H₂-receptors. Consequently, histamine released from mast cells and basophils during the early-phase allergic reaction may act as an important stimulatory signal for the initiation of the allergic inflammatory late-phase reaction by increasing local IL-5 production of allergen triggered T_H2 cells.     

Biological activity of recombinant murine interleukin-6 in interleukin-1 T cell assays

Interleukin-6 (also called B cell stimulatory factor 2, hepatocyte activating factor, interferon-β₂) has been shown to have effects on various lineages of hemopoietic cells. Some of its activities appear to overlap those of interleukin-1. In particular, recombinant murine IL-6 induced proliferation of phytohemagglutinin-activated thymocytes, an assay widely used to detect IL-1. In this report, we compared several features of IL-1 and IL-6 dependent thymocyte proliferation. The results indicate that IL-2 is the major second mediator of both IL-1 and IL-6 dependent proliferation. Finally, we tested whether IL-6 would also have activity in other T cell-based IL-1 assays using the T cell lymphoma LBRM33 1A5 and the T cell clone D10-G4.1. IL-6 had no activity in the latter two assays. These results indicate that IL-1 assays using LBRM33 1A5 and D10-G4.1 selectively detect Il-1, and are more specific assays for the detection of IL-1 in samples that may also contain IL-6.     

异烟腙类衍生物TJU103对人T细胞增殖和细胞因子分泌的影响

目的 :探讨异烟腙类衍生物TJU10 3体外对同种异体抗原刺激的T淋巴细胞增殖和细胞因子分泌的影响。方法 :采用MTT法和酶联免疫吸附实验 (ELISA)分别观察了TJU10 3对T细胞增殖和分泌细胞因子的影响。结果 :TJU10 3在 5 0mg/L浓度时 ,能够明显降低HLA半相合供、受者间混合淋巴细胞反应 (MLR)以及肿瘤坏死因子α(TNF α)和γ干扰素 (IFN γ)等细胞因子的分泌 (P <0 0 1)。结论 :TJU10 3能够特异性地与CD4 T细胞结合 ,降低CD4 T细胞的增殖和细胞因子分泌。     

Functional CD40 ligand expression on T lymphocytes in the absence of T cell receptor engagement: involvement in interleukin-2-induced interleukin-12 and interferon-γ production

Despite the fact that the great majority of T cells at the site of an inflammatory response are not antigen specific, the mechanisms leading to activation and recruitment of these bystander T cells are poorly understood. We previously reported that soluble (s)CD23 potentiated the interleukin (IL)-2-induced interferon (IFN)-γ production by T cells co-cultured with autologous monocytes in the absence of T cell receptor (TCR) engagement. Our present data demonstrate that the IL-2-induced IFN-γ secretion, in the presence but also in the absence of sCD23, is strictly IL-12 dependent, inasmuch as anti-IL-12 antibody abrogated both responses. Most interestingly, anti-CD40 ligand (CD40L) monoclonal antibody significantly inhibited IL-2-induced IL-12 as well as IFN-γ production. These results suggest that CD40L was expressed on T cells in the absence of TCR engagement. Indeed, purified unstimulated T cells readily expressed CD40L. IL-2 and monocytes did not up-regulate CD40L on resting T cells. It is proposed that low levels of CD40L expression on non-antigen stimulated T cells are sufficient to signal through CD40 molecules on accessory cells and to induce IL-12 secretion, which in turn can synergize with IL-2 for the induction of IFN-γ production, thus contributing to the inflammatory process.     

An analysis of the cellular requirements for the production of soluble interleukin-2 receptorsin vitro

Following activationin vitro, peripheral blood mononuclear cells (PBMC) express cell-associated interleukin-2 receptors (IL-2R) and also release soluble IL-2R into culture supernatants. The present studies were undertaken to define which normal cells were responsible for the release of soluble IL-2Rin vitro. Both cell-associated and soluble IL-2R were quantitatively measured with a sandwich enzyme-linked immunoassay employing two monoclonal antibodies. PBMC were separated into populations of surface immunoglobulin-negative cells (T cells and monocytes) and surface immunoglobulin-positive cells (B cells and monocytes), and the T-cell population was further separated into OKT4-positive (OKT4⁺) cells and OKT4-negative (OKT4^–) cells. Following activation with phytohemagglutinin, pokeweed mitogen, and the monoclonal antibody OKT3, large amounts of soluble IL-2R were released by PBMC, unseparated T cells, OKT4⁺ T cells, and OKT4^– T cells. The population containing B cells and monocytes made small but readily detectable amounts of soluble IL-2R when stimulated with these T-cell mitogens; likely the result of contaminating T cells in the population. However, when highly purified B cells were stimulated withStaphylococcus aureus Cowan and recombinant IL-2, they also released small amounts of soluble IL-2R. The release of soluble IL-2R by T cells appeared monocyte dependent when OKT3, but not phytohemagglutinin, was employed for activation, and monocytes themselves released no detectable IL-2R under the conditions employed. These studies define the cellular requirements for the release of soluble IL-2Rin vitro and demonstrate that such receptors are released by B cells, T cells, and both OKT4⁺ and OKT4^– T-cell subsets.     

淫羊藿甙对小鼠T淋巴细胞体外活化和增殖的影响

目的研究淫羊藿甙(ICA)对刀豆蛋白A(ConA)刺激的小鼠T淋巴细胞体外早期活化和增殖的影响。方法以MTT法检测T细胞药物毒性;利用流式细胞术(FCM)结合双色荧光抗体染色技术检测早期活化标志CD69分子的表达;运用流式细胞术结合活体染料羧基荧光素乙酰乙酸(CFDA-SE)染色技术和荧光抗体染色技术检测T细胞增殖。结果ICA在终浓度0.3、1.5、3.0μmol·L-1时对于CD69的表达以及淋巴细胞48和72h增殖,均有明显的抑制作用(P<0.01)。结论ICA能抑制ConA刺激的小鼠T淋巴细胞的早期活化和增殖,并呈剂量依赖性,即在最高浓度3.0μmol·L-1时抑制率最强。     

Role of LFA-1/ICAM-1 in interleukin-2-stimulated lymphocyte proliferation

Major adhesion routes between lymphoid cells involve the receptor/ligand pairs LFA-l/ICAM-1 and CD2/LFA-3, in addition to VLA or CD44 molecules. In this study we evaluated the role of these adhesion receptors in the proliferative response of lymphoid cells to interleukin-2 (IL-2). Blocking studies were performed with a panel of monoclonal antibodies (mAb) directed against these adhesion molecules. Selective inhibition of recombinant (r)IL-2-induced cell proliferation was observed with mAb directed against the a or /3 subunit of LFA-1 or to its ligand ICAM-1. Interestingly, rIL-2-induced proliferation was also inhibited by NKI-L16, an anti-la antibody known to enhance cell-cell interaction. Resting lymphocytes were preferentially susceptible to the inhibition, particularly in an early phase of culture and when stimulated with a relatively low dose of rIL-2. By using mAb that specifically could block distinct rIL-2 activation pathways, LFA-l/ICAM-1 interaction was found to be required for p55 IL-2 receptor (IL-2R)-mediated interaction of rIL-2 with its high-affinity receptor, but not for p75 IL-2R-mediated responses. Furthermore, it was shown that the rIL-2 response of T lymphocytes, but not of natural killer cells, was dependent on LFA-l/ICAM-1 interaction. This suggests that LFA-l/ICAM-1 interaction is required for an optimal rIL-2 response of cells capable of IL-2 secretion. Our data provide evidence for the hypothesis that adhesion receptor-directed release of IL-2 may result in a locally high concentration of IL-2 that triggers high-affinity IL-2R signaling and up-regulates p55 IL-2R to enhance cytokine responsiveness.     

Regulation of 4-1BB expression by cell-cell interactions and the cytokines,interleukin-2 and interleukin-4

4-1BB expression increased gradually following T cell activation, and by day 3 post-stimulation with immobilized anti-CD3 (anti-CD3i) or concanavalin A (Con A), splenic T cells were routinely 35–45% 4-1BB⁺ by flow cytometric analysis. 4-1BB was expressed on activated CD8⁺, CD4⁺, CD28⁺ and CD45RB⁺ T cells. Optimal 4-1BB expression was seen by day 6 post-stimulation and was cell density dependent. When T cells were cultured for 6 days at 1 × 10⁶/well in a 24-well plate with anti-CD3i, 82% of the cells were 4-1BB⁺. In contrast, at lower cell densities (4 × 10⁵, 2 × 10⁵ and 1 × 10⁵), optimal 4-1BB expression was observed only if the cultures were supplemented with recombinant interleukin-2 (IL-2) or recombinant IL-4 (IL-4). In agreement, with these results, modes of inducing endogenous IL-2 production such as cross-linking the costimulatory molecule, CD28, or the addition of syngeneic accessory cells to T cells activated with anti-CD3i, resulted in high levels of 4-1BB expression. The addition of interleukin-1α(IL-1α) or interferon-γ (IFN-γ) did not increase 4-1BB expression on anti-CD3i-activated T cells. In addition, if T cells were incubated with IL-2, IL-4, IL-1α, IFN-γ or anti-CD28 alone, no 4-1BB expression was induced. T cells activated with soluble anti-CD3 (anti-CD3s) in the presence of IL-2, IL-4, or accessory cells, did not express higher levels of 4-1BB on their cell surface. These data suggest that initial events crucial for efficient T cell activation, such as signals delivered through the T cell receptor/CD3 complex and the CD28 molecule, are instrumental in regulating subsequent 4-1BB expression.