自身免疫性甲状腺疾病外周血淋巴细胞体外分泌IgG及抑制功能的研究

本实验在体外测定了自身免疫性甲状腺疾病(autoimmune thyroid disease,AITD)患者外周血淋巴细胞培养的上清IgG含量,并根据Concanaralin(ConA)可诱导抑制性T细胞(suppressor T cell,Ts)的原理,检测了ConA诱导Ts对淋巴细胞分泌IgG的抑制率(suppressiverate,SR)。结果表明AITD患者淋巴细胞分泌IgG与正常人无明显差别(P>0.05),而其SR较正常对照显著低下(P<0.05),且SR与Graves病患者血T_3值呈负相关。实验进一步证明甲状腺素在体外并不影响ConA诱导Ts的抑制作用。本研究结果支持本病具有Ts功能的缺陷,且提示这种异常并不是继发于本病,而是本病原发性的免疫表现。     

慢性乙型肝炎患者外周血T淋巴细胞亚群的变化

目的了解慢性乙型肝炎患者的细胞免疫功能. 方法用免疫花环法测定41例慢性乙肝患者外周血T细胞亚群并分析其变化. 结果 CD3、CD4细胞以及CD4/CD8比值较正常人显著减少(p<0.01),CD8细胞较正常人明显升高(p<0.01). 结论慢性乙肝患者外周血T细胞亚群紊乱,免疫功能低下.     

再生障碍性贫血患者外周血T细胞亚群和T细胞受体表达水平及其意义

采用流式细胞仪及间接免疫荧光法检测30例再生障碍性贫血患者外周血中T细胞亚群及T细胞表面受体表达水平,并与健康对照组相比,结果表明:70％再障患者存在CD4/CD8比例倒置及CD8~+％异常增高;50％再障患者外周血γδ-T细胞亚群及其在T淋巴细胞总体中所占比例均显著增高;而αβT细胞亚群及TirA~+细胞百分率与正常对照组相比无显著性差异。提示:半数以上再障患者外周血中存在异常增多的γδT细胞及Ts细胞亚群。并可通过其直接或间接作用抑制造血,从而导致再障的发生。     

肝素诱导的DC对慢性乙肝患者Th1/Th2分化的影响

探讨肝素体外对慢性乙肝患者CD4~+T细胞亚群分化的影响。分离慢性乙肝患者外周血单核细胞,以rhIL-4(50 ng/ml)、rhGM-CSF(10 ng/ml)、rhTNF-α(100 u/ml)和肝素(50 u/ml)诱导培养DC。免疫磁珠分离外周血CD4~+T细胞亚群,ELISA法检测DC或CD4~+T细胞培养上清中IL-6、IL-12、IFN-γ和IL-4的含量。以流式细胞仪检测肝素处理前后DC表面CD1a、CD83、CD80、CD86、HLA-DR分子表达情况。结果:纯化的慢性乙肝患者CD4~+T细胞培养上清中IFN-γ显著低于同期培养的正常人CD4~+T细胞(P0.05);肝素处理的慢性乙型肝炎患者的DC表达CD1a、CD83、CD80、CD86、HLA-DR分子水平及分泌的IL-12水平明显升高(分别P0.05、P0.01);而分泌的IL-6水平降低(P0.05)。肝素处理后的DC与慢性乙肝患者外周血中CD4~+T细胞共培养后,其上清中Th1型细胞因子IFN-γ的水平明显升高(P0.01)。体外肝素处理后的慢性乙肝患者的DC能部分改善外周血Th1细胞分化的不足。     

T淋巴细胞与结核免疫

关于T 细胞与结核免疫,近年来的研究表明,CD_4~+、CD_8~+T 细胞都参与抗结核的保护作用。CD_8~+ T 细胞的增殖和分化需要由CD_4~+T 细胞提供的IL-2。CD_4~+T 细胞可通过淋巴因子的分泌激活巨噬细胞,CD_4~+T 细胞还可过继转移DTH 反应,并能激活体内的抗结核防御机制。接受过BCG 预防注射的健康人的外周血单核细胞,在体外用BCG 刺激5天后可诱导产生抑制细胞,这种抑制细胞的抑制作用是特异性的,并表现为MHC 制约性,用单克隆抗体检测发现它是CD_4~+亚类。     

人体T细胞CD_8~+VV~+亚群在活动性SLE患者中显示反抑制活性

反抑制T 细胞(TCS)借助T 辅助细胞对抗T 抑制细胞的抑制效应,参与免疫调节。小鼠和人体的TCS 都表达长绒毛野豌豆凝集素受体(VV—R)。人体TCS 则主要是CD_8~+VV~+T 亚群,和临床多种疾病免疫功能异常密切相关。本文用抗VV 抗体单色和双色免疫荧光法和流式法细胞术(FCM)测知正常人VV~+T 亚群的比例是16.6±2.9%。用FCM 双色免疫荧光法检测到活动期SLE 患者CD_8VV~+亚群百分比高达23.63%和对照组(8.50%)及稳定期(14.49%)相比差异显著。然而CD_4~+VV~+亚群在三组SLE 中基本不变。进一步深入分析是做有关CD_8VV~+T 细胞亚群的功能测定,证实活动性SLE 患者VV~+T 细胞具有比正常个体高得多的反抑制活性,相对反应值从100%提高到485%和625%。提示CD_(?)VV~+细胞和SLE 免疫调节功能紊乱密切相关。     

纳洛酮对人体短时剧烈运动中机体免疫功能的影响

本文就纳洛酮对击剑运动员短时间剧烈运动前后机体免疫功能的影响进行了初步探讨,结果表明:运动前静脉注射纳洛酮后跑步4000米,外周血T细胞亚群的变化表现为CD_8~+细胞亚群百分率升高,CD_4~+/CD_8~+比值下降;运动后外周血白细胞诱生干扰素的能力显著高于运动前,P值<0.01。运动后NK细胞活性及白细胞吞噬百分率有下降的趋势,但吞噬指数显著增高。纳洛酮对外周血白细胞诱生干扰素的能力、NK细胞活性及白细胞吞噬功能均未见有明显影响。     

慢性乙型肝炎患者外周血T淋巴细胞亚群的变化

目的：了解慢性乙型肝炎患者的细胞免疫功能。方法：用免疫花环法测定41例慢乙肝患者外周血T细胞亚群并分析其变化。结果：CD3、CD4细胞以及CD4／CD8比值较正常人显著减少（p＜0．01），CD8细胞较正常人明显升高（p＜0．01）。结论：慢性乙肝患者外周血T细胞亚群紊乱，免疫功能低下。     

慢性粒细胞白血病患者外周血高表达CD266~+Foxp3~+γδ~+T细胞亚群

目的分析慢性粒细胞白血病慢性期(CML-CP)患者外周血γδT细胞及其亚群的表达情况,及其与临床一线酪氨酸激酶抑制剂(TKI)药物治疗疗效的相关性。探讨成纤维细胞生长因子诱导因子14(Fn14,又称CD266)在γδT细胞功能亚群中的表达情况及其与临床疗效的相关性。方法采用流式细胞术检测CML-CP患者以及正常对照组外周血γδT细胞及其功能亚群的表达情况,并探讨γδT细胞各亚群表达比例与临床一线TKI药物治疗疗效的相关性。结果 CML-CP患者外周血γδT细胞功能亚群CD266~+γδ~+T细胞、Foxp3~+γδ+T细胞、CD266+Foxp3~+γδ~+T细胞、CD266~+Vδ1~+T细胞、Foxp3~+Vδ1~+T细胞、CD266~+Foxp3~+Vδ1~+T细胞、CD266~+Vδ2~+T细胞、Foxp3+Vδ2+T细胞和CD266~+Foxp3~+Vδ2~+T细胞的表达比例均显著高于正常对照组。经过一线TKI药物治疗后达到完全血液学反应的基础上,达到警告治疗反应或治疗失败的CML-CP患者外周血CD266~+Foxp3~+Vδ2~+T细胞亚群比例显著增高。Logistic回归分析亦显示CD266~+Foxp3~+Vδ2~+T细胞亚群的表达比例为CML-CP患者经过一线TKI药物治疗后发生难治的危险因素。结论 CD266~+Foxp3~+γδT细胞亚群与CML-CP患者的临床一线TKI药物治疗疗效密切相关。Fn14(CD266)信号通路可能成为复发难治CML患者的新的治疗靶点。     

类风温性关节炎患者T抑制诱导细胞亚群缺陷

类风湿性关节炎(RA)存在的T细胞调节功能障碍已得到公认,但对T辅助细胞(CD_4)/T抑制细胞(CD_8)比值的研究尚无一致的结论。晚近,单克隆抗体的研究已将CD_4淋巴细胞分为CD_4~+ 4B4~+和CD4~+2H4~+两种亚群,前者为辅助诱导细胞(HI),后者为抑制诱导细胞(SI)。本文作者采用双色免疫荧光及流式细胞测定技术,以抗4B4、2H4、CD4和CD8抗体对RA及其它关节病(OAD)进行了T细胞亚群测定。结果表明,RA患     

Effect of phorbol myristate acetate on T cell colony formation, interleukin-2 (IL-2) receptor expression and IL-2 production by cells from patients at all stages of HIV infection.

We and others have shown that several T cell responses induced by the mitogen phytohaemagglutinin (PHA), including T cell colony formation, IL-2 receptor (IL-2R) expression, and IL-2 production are impaired in patients with AIDS and lymphadenopathy syndrome (LAS). We investigated whether phorbol myristate acetate (PMA) could act in synergy with PHA (as it does in healthy subjects) to enhance in vitro T cell responses of patients at all stages of infection by HIV. In AIDS patients with opportunistic infections (AIDS/OI), PHA + IL-2 + PMA led to a total disappearance of T cell colonies in 10/11 patients, among whom six already displayed very low numbers of colonies induced by PHA + IL-2 (less than 50 colonies/5 x 10(4) cells). In contrast, T cell colony formation induced by PHA + IL-2 + PMA was maintained or increased, compared with that induced by PHA + IL-2, in five out of six AIDS patients with Kaposi's sarcoma (AIDS/KS), 10/14 LAS and six out of seven HIV-seropositive asymptomatic (HIV+/AS) homosexuals. In these three groups of patients, a low percentage of colony cells induced by PHA + IL-2 + PMA expressed CD3 and CD4 molecules, but 50-89% of cells were IL-2R (Tac) positive, as in healthy controls. Studies on T cell activation and IL-2 production were performed on a selected group of 12 HIV-infected patients for whom sufficient numbers of lymphocytes could be obtained. PMA induced CD4 down-modulation in controls and in HIV-infected patients. However, CD3 down-modulation and induction of the Tac chain of IL-2R by PMA were significantly impaired in patients, compared with controls, and these two parameters were correlated. Although PHA alone induced virtually normal levels of Tac antigen on patients' cells, Tac induction by PHA + PMA was significantly decreased in patients versus controls. Cells from five out of 10 patients tested failed to produce detectable amounts of IL-2 after PHA stimulation, whereas IL-2 production increased significantly in all patients tested (n = 9) after PHA + PMA, with a level of IL-2 activity significantly higher than in controls. No correlation was found in this group of patients between the effects of PMA + PHA on T cell colony formation, Tac expression, or IL-2 production, as compared with PHA alone. Taken together, our results indicate that in vitro T cell functional studies with PMA may be useful to evaluate better the defects of T cell activation in HIV-infected patients.     

Immune activation and T cell subset abnormalities in circulation of patients with recently diagnosed type I diabetes mellitus.

The peripheral blood lymphocytes from patients with insulin-dependent diabetes mellitus (IDDM) and healthy controls were analysed for the HLA-DR+, interleukin-2 receptor-positive (IL-2R+) activating antigens, and for CD45R+ and CDw29+ subsets from the purified CD4+ and CD8+ T cells populations. Patients with IDDM had an increased percentage of HLA-DR+ and IL-2R+ cells in both CD4+ and CD8+ T cells. However, the percentage of CD4+ CD45R+ suppressor/inducer T cells were decreased and CD4+ CDw29+ helper/inducer T cells increased in all patients with IDDM, compared with healthy controls. Thus, IDDM patients exhibit a deficiency in the CD4+ CD45R+ suppressor/inducer T cell subsets, which is probably related to the autoimmune phenomenon in this disease. In contrast, the percentage of CD8+ CDw29+ T cell subsets showed no major differences between patients with IDDM and controls. An alteration in the CD4+ CD45R+ and CD4+ CDw29+ T cell subsets appears to be a characteristic feature, and may relate to the impaired cell-mediated immunity in IDDM. These data provide new evidence for T cell dysregulation in IDDM.     

Establishment of stable CD8+ suppressor T cell clones and the analysis of their suppressive function.

Stable CD8+ suppressor T cell (Ts) clones were established by a relatively simple method. Keyhole limpet hemocyanin (KLH)-primed spleen cells from C3H mice were depleted of B cells and CD4+ T cells by panning and cytotoxic treatment, and the resulting CD8+ T cells were periodically stimulated with antigen and irradiated syngeneic spleen cells followed by manifestation in interleukin-2 (IL-2) containing medium. T cell clones with a definite suppressor function were established by limiting dilution. They were defined as classical effector type Ts of CD8+ phenotype as they had constant and definite suppressor functions in antigen-induced T cell proliferation and specific antibody response against T cell-dependent antigens without detectable cytotoxic activity against both antigen presenting cells (APC) and helper T cells (Th). They showed no helper activity for B cells and produced no detectable helper type lymphokines such as IL-2 and IL-4. CD8+ Ts clones were able to inhibit the antigen-induced IL-2 production of normal and cloned T cells. Their suppressive activity was antigen-nonspecific and major histocompatibility complex-unrestricted. CD8+ Ts clones were also able to suppress the proliferative response of Th clones induced by immobilized anti-T cell receptor (TcR) and anti-CD3 mAbs but not the response induced by concanavalin A (ConA) and IL-2. All the CD8+ T cell clones established independently utilized the TcR V beta 8 gene. Syngeneic antigen presenting cells could induce proliferation of these CD8+ clones, which was blocked by anti-CD8 and anti-I-Ak monoclonal antibody (mAb) but not by anti-class I mAbs. The stimulation of CD8+ Ts clones with immobilized anti-CD3 resulted in the release of a suppressor factor(s) that potently inhibited the antigen-induced proliferation of CD4+ Th clones and the in vitro secondary antibody formation.     

IL-24对非小细胞肺癌患者CD8+T细胞体外功能的影响

目的观察IL-24在体外对非小细胞肺癌(non-small cell lung cancer,NSCLC)患者CD8⁺T细胞功能的影响。方法本研究入组28例NSCLC患者和17例健康对照者,收集外周血单个核细胞(peripheral blood mononuclear cells,PBMC)和支气管肺泡灌洗液(bronchoalveolar lavage fluid,BALF),分选CD8⁺T细胞,反转录实时定量PCR检测CD8⁺T细胞中IL-24受体(IL-20R1、IL-20R2和IL-22R1)mRNA的相对表达量。不同浓度重组人IL-24(10 ng/ml和100 ng/ml)刺激纯化CD8⁺T细胞后,流式细胞术检测穿孔素和颗粒酶B的表达变化。建立CD8⁺T细胞和NSCLC细胞系NCI-H1882细胞的直接接触和间接接触体外共培养系统,观察IL-24刺激后CD8⁺T细胞诱导靶细胞死亡比例,以及IFN-γ和TNF-α的表达变化。组间比较采用t检验或LSD-t检验。结果CD8⁺T细胞中未检测到IL-22R1 mRNA表达,CD8⁺T细胞中IL-20R1和IL-20R2 mRNA相对表达量在健康对照者和NSCLC患者之间以及在非肿瘤部位和肿瘤部位之间的差异均无统计学意义(P>0.05)。NSCLC患者外周血和肿瘤部位CD8⁺T细胞中穿孔素和颗粒酶B水平显著低于健康对照者和非肿瘤部位(P<0.05),低浓度IL-24(10 ng/ml)刺激不影响CD8⁺T细胞中穿孔素和颗粒酶B水平(P>0.05),而高浓度IL-24(100 ng/ml)则显著提升NSCLC患者CD8⁺T细胞中穿孔素和颗粒酶B水平(P<0.05)。直接接触共培养系统中,高浓度IL-24(100 ng/ml)刺激NSCLC患者肿瘤部位CD8⁺T细胞后可诱导靶细胞死亡比例升高,以及IFN-γ和TNF-α表达升高,而低浓度IL-24(10 ng/ml)刺激对CD8⁺T细胞诱导的靶细胞死亡和细胞因子分泌无显著影响。间接接触共培养系统中,IL-24刺激对CD8⁺T细胞诱导的靶细胞死亡和细胞因子分泌均无显著影响。结论高浓度IL-24在体外可增强NSCLC患者CD8⁺T细胞的直接细胞杀伤功能,但在体内IL-24可能并不影响CD8⁺T细胞的功能。     

晚期肺癌患者调节性T细胞和IL-10表达的变化

探讨晚期肺癌患者的CD4＋CD25＋调节性T细胞（Treg）、IL-10及其他T细胞亚群表达的临床意义。检测晚期肺癌患者92例和正常对照者64名的IL-10（ELISA法）、CD4＋CD25＋调节性T细胞及其他T细胞亚群（流式细胞仪法）。结果显示：肺癌组血清IL-10明显大于对照组（278±34ng/L vs 122±65ng/L,P〈0.01）、CD4＋CD25＋Treg细胞明显大于对照组[（17.8±5.2）%vs（7.1±0.4）%,P〈0.01]、CD3＋、CD4＋、CD8＋CD28＋和NK细胞明显小于对照组[（58.4±7.8）%vs（78.2±6.4）%,P≤0.05;（34.4±7.6）%vs（44.9±8.4）%,P〈0.01;（9.4±3.6）%vs（16.5±2.7）%,P〈0.01;（9.4±3.6）%vs（18.5±7.2）%,P〈0.01]。CD8＋T细胞明显高于对照组[（37.8±6.5）%vs（31.8±5.1）%,P〈0.01]。CD4＋CD25＋Treg细胞、IL-10与CD8＋CD28＋细胞、NK细胞呈明显负相关。这些结果表明,CD4＋CD25＋Treg细胞与IL-10增多为晚期肺癌患者免疫功能受损的表现。     

T lymphocyte subsets, suppressor and contrasuppressor cell functions, and production of interleukin-2 in the peripheral blood of rheumatic fever patients and their apparently healthy siblings

We studied CD4 and CD8 T cell subsets, suppressor cell function, production of IL-2, and immune contrasuppressor cell activity in 21 patients with rheumatic fever (RF), both at the time of their first acute episode and 3 months later (recovery phase). As controls we studied their healthy sibling nearest in age, as well as age- and sex-matched unrelated normal subjects. In the acute phase we found CD4+ T cells to be high, concanavalin A-induced suppression to be low, and production of IL-2 to be significantly decreased, as compared to the normal unrelated controls. The addition of contrasuppressor cells (VV+) to cell cocultures resulted in an increase in proliferation by mononuclear cells (MNC) in response to streptococcal M antigen but not to C carbohydrate antigen. In the recovery phase, CD4+ T cells became normal, CD8+ T cells rose above normal, and the suppressor cell functions (concanavalin-A-induced and spontaneously expanded), as well as the production of IL-2, fell further. Siblings were found to have increased CD8+ T cells and decreased production of IL-2, as compared to the unrelated controls. These findings indicate that important immunoregulatory disturbances occur during the acute phase of rheumatic fever, some of which persist, accentuate, or change during the recovery phase. The findings in siblings could be related either to streptococcal infection or to a familial immunoregulatory aberration.     

Immunoregulatory CD4+ CD45R+ suppressor/inducer T lymphocyte subsets and impaired cell-mediated immunity in patients with Down''s syndrome.

The monoclonal antibodies 2H4 and 4B4 allow CD4+ and CD8+ T lymphocytes to be subdivided into CD45R+ and CDW29+ functional subpopulations. The CD4+ CD45R+ lymphocytes are designated as suppressor/inducer and CD4+ CDW29+ as helper/inducer subsets. Peripheral blood lymphocytes from 19 patients with Down's syndrome and 19 age- and sex-matched normal controls were analysed for the CD45R+ and CDW29+ subsets from the CD4+ and CD8+ T lymphocytes. The percentage of CD4+ CD45R+ cells (suppressor inducer) was markedly increased and of CD4+ CDW29+ cells (helper/inducer) decreased in all patients with Down's syndrome. In contract, the percentage of CD8+ CD45R+ and CD8+ CDW29+ subsets showed no major differences between patients with Down's syndrome and normal controls. Moreover, an alteration in the CD4+ and CD45R+ and CD4+ CDW29+ T cell subsets was accompanied by a markedly reduced proliferative response to phytohaemagglutinin and concanavalin A stimulation of the CD4+ T lymphocytes. Thus, a deficiency exists in patients with Down's syndrome in the CD4+ CDW29+ helper/inducer T cell subset which may contribute to their impaired cell-mediated immunity.     

天花粉蛋白合成肽通过激活抑制性CD8~+ T细胞诱导免疫抑制

为了探讨天花粉蛋白合成肽(M-Tk)诱导免疫抑制的机制,应用小鼠针对可溶性抗原OVA的增殖系统,检测M-Tk处理后对淋巴细胞增殖的抑制作用和细胞因子格局的变化,以及M-Tk激活的T淋巴细胞亚群的表型及功能。结果表明,M-Tk可诱导一群CD8+ T抑制性细胞,其表型为CD8+CD28-CTLA4+,采用双层非接触共培养系统发现,该群抑制性细胞主要依赖于细胞因子IL-10、IL-4和细胞间接触发挥免疫抑制作用。     

Defective integration of activating signals derived from the T cell receptor (TCR) and costimulatory molecules in both CD4+ and CD8+ T lymphocytes of common variable immunodeficiency (CVID) patients

CVID is characterized by hypogammaglobulinaemia and impaired antibody production. Previous studies demonstrated defects at the T cell level. In the present study the response of purified CD4⁺ and CD8⁺ T lymphocytes to stimulation with anti-TCR monoclonal antibody (the first signal) in combination with anti-CD4 or anti-CD8, anti-CD2 and anti-CD28 MoAbs (the costimulatory signals) was investigated. Both CD4⁺ and CD8⁺ T cells from the patients showed significantly reduced IL-2 release following stimulation via TCR and costimulation via CD4 or CD8 and CD2, respectively. However, normal IL-2 production following TCR plus phorbol myristate acetate (PMA) costimulation and normal expression of an early activation marker, CD69, after TCR + CD28 stimulation indicated that TCR was able to transduce a signal. Furthermore, both IL-2 and IL-4 release were impaired in CD4⁺ lymphocytes following TCR + CD28 stimulation. In addition, stimulation via TCR + CD28 resulted in significantly decreased expression of CD40 ligand in the patients. These results suggest that the integration of activating signals derived from the TCR and costimulatory molecules is defective in CVID patients; the defect is not confined to costimulation via a single molecule, or restricted to cells producing Thl-type cytokines such as IL-2, and is expressed in both CD4⁺ and CD8⁺T cell subsets.     

Detection of circulating parasite antigen and specific antibody in Toxocara canis infections.

IL-2 production by peripheral blood mononuclear cells (PBM) is decreased in patients with systemic lupus erythematosus (SLE). This defect can be reversed by the removal of CD8+ lymphocytes. The purpose of these studies was to determine whether the CD8+ IL-2 suppressor cells comprise a specific subset or whether all CD8+ cells have this activity. Lymphocyte subsets were identified and separated by two-colour flow cytometry prior to a 48 h mitogen stimulation. The CD8+ cells that suppressed IL-2 production co-expressed HLA DR and were radiosensitive. Other markers co-expressed by CD8+ cells which are found on suppressor cells such as Leu 15 (CD11), Leu 11 (CD16), and Leu 7 were also found on the CD8+ IL-2 suppressor cell population in SLE. In healthy subjects, removal of CD16+, but not of CD8+ cells markedly elevated the production of IL-2. The CD8- CD16+ non-T cell subset suppressed IL-2 production by normal and SLE PBM in autologous and allogeneic combinations. This subset may be a human equivalent of the murine natural suppressor cells. These results demonstrate that the cells that suppress IL-2 production in SLE are heterogeneous, and suggest that they belong to more than one lineage.