系统感染白色念珠菌的Balb/c小鼠免疫功能探讨

目的 探讨Balb/c小鼠系统感染白色念珠菌后免疫功能的动态变化. 方法 MTT法检测ConA诱导的T淋巴细胞增殖功能;ELISA法测定血清IL-18 、IFN-γ和IL-4水平;RBC-C3bRR法观察红细胞的黏附功能;真菌培养计算左肾白色念珠菌数.结果 高低剂量组感染组T淋巴细胞增殖功能呈上升趋势,至感染后第 7天达正常水平;IL-18水平均从感染第1天骤降后,至感染后第7天达正常水平;而IFN-γ始终高于正常对照组及免疫功能低下组;IL-4于感染1 d后降低,至第7天后高于正常对照组.低剂量组感染白色念珠菌后对RBC-C3bRR有刺激增强作用 .高低剂量组的白色念珠菌的廓清能力随时间延长均增强.结论 Balb/c 小鼠高低剂量感染白色念珠菌时,对ConA诱导的T淋巴细胞增殖功能有一定的刺激增强作用;感染早期多以Th1 型细胞应答为主,后期Th1/Th2型细胞因子均升高,示体液免疫也参与了机体抗白色念珠菌感染.Blab/c小鼠低剂量感染白色念珠菌对RBC-C3bRR有刺激增强作用.Blab/c小鼠高低剂量感染白色念珠菌后,其廓清能力随时间延长而不断增强.     

rHu TNF-α促进人胚和小鼠胸腺细胞增殖作用的研究

本文观察了rHuTNF-α对ConA诱导的10～15日龄BALB/C乳鼠胸腺细胞、脾细胞以及PHA或TPA诱导的20～32周龄人胚胸腺细胞增殖的调节作用。结果表明:rHuTNF-a对上述增殖作用均有显著的促进作用,并且呈剂量依赖关系;rHuTNF-a对rHuIL-2促进ConA诱导的小鼠胸腺细胞和脾细胞的增殖反应有协同作用。此结果提示rHuTNF-a在促进丝裂原诱导的胸腺细胞增殖作用中无明显种属特异性,并且对于了解TNF-a在胸腺细胞发育中的作用以及在某些病毒性感染疾病患者血清中高水平的TNF-a活性与胸腺功能改变的关系方面均有一定的意义。     

HFRSV感染乳鼠的脾细胞增殖及IL-2产生能力的变化

本文研究了HFRSV感染3～5日龄BALB/C新生乳鼠脾脏的病理变化、脾细胞对丝裂原或/和细胞因子诱导的增殖效应及IL-2产生能力的变化。结果表明:感染组鼠较正常组鼠(1)脾脏体积明显缩小,脾细胞数量显著减少;(2)脾细胞在无刺激原条件下培养,~3H-TdR掺入cpm值明显升高;(3)脾细胞对ConA或/和rHuTNF-α、rHu IL-2诱导的增殖反应显著降低,但是rHu IL-2、rHuTNF-α对ConA诱导的感染鼠脾细胞增殖反应仍有明显的促进作用,且这两种细胞因子有协同效应;(4)脾细胞经ConA诱导后的IL-2产生能力显著降低。此结果对于进一步了解HFRSV感染过程中细胞免疫功能变化有重要意义,同时也为临床应用细胞因子进行免疫治疗提供了有价值的实验依据。     

雷公藤内酯醇抗移植排斥反应的研究

应用小鼠同种异位心肌移植模型,证实雷公藤内酯醇(triptolide,Tri)可明显延长移植心的存活期。对T淋巴细胞增殖活性影响的实验结果表明,Tri对ConA激活的淋巴细胞转化及特异性抗原激活的混合淋巴细胞反应均有明显地抑制作用;同时,Tri对ConA诱导的IL2产生无影响,但对IL2受体的表达有明显地抑制作用。     

骨髓源间充质干细胞在异基因小鼠免疫器官内的分布及其免疫功能调节作用

目的：探讨骨髓源间充质干细胞(Bone marrow mesenchymal stem cells，BMSC)在异基因小鼠免疫器官内的分布及其免疫调节作用。方法：以CM-Dil荧光染料示踪BMSC的体内分布情况，并辅以PCR检测Y染色体的方法进一步鉴定；体外实验采用MTT法、ELISA和FACS等方法检测BMSC的免疫调节作用。结果：BMSC可进入并较长期(30天)存在于异基因小鼠免疫器官内；在体外，BALB／C小鼠的BMSC对由ConA诱导的BALB／C和C57BL／6(B6)和BXSB小鼠的T细胞增殖均有抑制作用；而对前两种小鼠由12S诱导的B细胞增殖和分泌k方面表现为促进作用，对BXSB小鼠由IPS诱导的B细胞增殖和k分泌有抑制作用。BALB／C小鼠的BMSC对BALB／C和B6小鼠由ConA诱导的IL-4生成细胞的数量无明显影响，却可降低由ConA诱导的两种品系小鼠的IFN-γ生成细胞的数量；但对于BXSB小鼠却不同，BALB／C的BMSC可降低由ConA诱导的BXSB小鼠的IL-4生成细胞的数量，而提高由ConA诱导的IFN-γ生成细胞的数量。结论：异基因BMSC不但可进入受体的免疫器官，且可较长期(30天)存在；另外，BMSC对同基因正常、异基因正常和异基因自身免疫病的个体均有一定程度的免疫调节作用。     

心房肽对自发性高血压大鼠免疫细胞增殖活性的影响

本文探讨了心房肽对免疫细胞增殖活性的影响。结果表明,心房肽可以直接促进大鼠胸腺细胞的增殖反应,亦可协同ConA促进脾细胞及T细胞的增殖反应,协同IL-2促进经ConA诱导的淋巴母细胞增殖;心房肽还可促进ConA诱导的大鼠脾细胞产生IL-2。但心房肽并不影响B细胞的增殖反应,也不影响巨噬细胞产生IL-1。同时发现,自发性高血压大鼠脾细胞和胸腺细胞对心房肽与ConA协同刺激的增殖反应性低下。     

血吸虫感染宿主免疫抑制现象的研究 Ⅱ.日本血吸虫感染鼠血清的免疫抑制作用

在正常鼠脾淋巴细胞和刀豆素A(ConA)的培养物中,加入感染日本血吸虫的同系鼠血清,发现血清浓度在2.5,5和10％时,对ConA诱导的增生应答均有抑制作用,进一步实验是在脾淋巴细胞和ConA的培养物中加入10％的感染后1～10周的鼠血清,以观察对脾淋巴细胞增生应答的抑制作用有无显著变化。结果表明,正常鼠脾淋巴细胞对ConA的应答,加入感染后第5周血清时,其抑制作用明显减弱。然而,感染鼠脾淋巴细胞对ConA的增生应答,加入感染后第5周血清时,其抑制作用的减弱不明显。提示感染血清对ConA增生应答的免疫抑制作用可能受双重因素影响,包括感染后某时期血清本身抑制作用的强弱,以及这一时期的淋巴细胞对ConA应答能力大小。     

生脉注射液对小鼠免疫功能的影响

探讨生脉注射液对小鼠免疫功能的作用。应用单克隆抗体和放射性掺入法检测胸腺T细胞和ConA诱导的脾细胞增殖反应等技术 ,测定生脉注射液对小鼠免疫功能影响的指标。结果表明 :生脉注射液增加胸腺Thy 1+细胞百分率 ,明显增强ConA诱导的脾细胞增殖 ,增加腹腔巨噬细胞的吞噬功能 ,提高外周血T淋巴细胞酯酶染色率 ,使血清IgG含量增加 ,明显增加胸腺、脾脏的重量。因而生脉注射液具有提高小鼠免疫功能的作用 ,为一免疫增强剂     

超抗原SEB活化的NKT细胞亚群及耐受功能的研究

目的 研究多肽类抗原.超抗原金黄色葡萄球菌肠毒索B(staphylococcal entcrotoxin B,SEB)活化的NKT细胞亚群及耐受特征.方法 小鼠脾细胞分别与SEB和ConA体外培养,MTT方法 测定细胞增殖;特异性耐受特征的研究使用体外活化第3天的细胞,吸出上清后添加二次抗原ConA、LPS、IL-2,继续培养3 d,MTT方法测定细胞对二次抗原的应答反应能力.用流式细胞测定法解析SEB和ConA体外活化的淋巴细胞在第0、5、10和15天时的T和NKT淋巴细胞亚群.结果 SEB和ConA均诱导了小鼠淋巴细胞在体外增殖;SEB活化的淋巴细胞对ConA、LPS和IL-2的二次应答反应能力消失,细胞增殖的OD570nm值由一次应答反应的0.433±0.07分别下降到0.19±0.01、0.14 ±0.02和0.15±0.04(P＜0.01).EonA活化的淋巴细胞的二次应答反应依然存在并依赖IL-2.SEB活化的淋巴细胞是CD4+NK1.1+和CD8+NK1.1+NKT细胞,而不是CD4-CD8-NKT细胞.ConA活化的淋巴细胞是CD3+、CD4+和CD8+T淋巴细胞并包括了CD4-C08-/CD3+NK1.1+NKT细胞.结论 除了α-Calcer脂多糖类抗原能够活化NKT细胞,多肽类超抗原SEB也能活化NKT细胞并具有特异性免疫耐受功能.能够介导免疫耐受的NKT细胞亚群可能是SEB活化的CIM+、CD8+NKT细胞而不是CD4-CD8-/CD3+NK1.1+NKT细胞.     

超抗原SEB诱导的免疫耐受与细胞因子无关

目的 研究超抗原金黄色葡萄球菌肠毒素B(staphylococcal enterotoxin B,SEB)诱导的耐受效应和耐受调节机制.方法 收集SEB活化10天的细胞,充分洗涤后做为效应细胞,分别与刀豆蛋白(ConA)、脂多糖(LPS)和白介素-2(IL-2)共同培养,用MTT方法测定细胞的耐受性应答反应.正常淋巴细胞在与ConA、LPS、IL-2共同培养的同时添加效应细胞,用MTF方法测定效应细胞的抑制性应答反应功能.流式细胞技术解析耐受性效应细胞类型.结果 效应细胞对ConA、LPS和IL-2的应答反应能力明显降低(P<0.01,n=3),但保持对ConA的应答反应能力.效应细胞抑制正常淋巴细胞与ConA、LPS和IL-2的应答反应(P<0.01,n=3),尤其抑制ConA和IL-2诱导的细胞增殖.SEB活化的效应细胞中CD8+NK1.1+、TcRVβ8+NK1.1+和CD4+NK1.1+NKT细胞以及TcRVβ8+T、CD8+T细胞数量明显增加(P<0.01和P<0.05,n=4).结论 超抗原SEB诱导的耐受性应答反应是效应细胞的直接作用,与细胞因子无关;这些效应细胞能抑制T淋巴细胞增殖,并保存识别ConA的受体功能.     

Effect of Systemic Candidiasis on Blastogenesis of Lymphocytes from Germfree and Conventional Rats

Germfree and conventional rats were challenged (intravenously) with Candida albicans and sacrificed at various times after infection, and their spleen cells were harvested to examine the effect of disseminated candidiasis on in vitro lymphocyte hypersensitivity to Candida antigens (CA). Results showed that conventional rat splenocytes, initially responsive in vitro to stimulation by CA, manifested a depression in CA-specific responsiveness after challenge with viable C. albicans (days 3 to 6 postchallenge). In addition, the latter splenocyte response to phytohemagglutinin (PHA) and concanavalin A (ConA) was suppressed by 3 to 6 days after challenge with Candida. In contrast to conventional rats, the response of germfree rat splenocytes to CA was insignificant before challenge with C. albicans, and it was increased at 9 days after infection. The response of uninfected germfree rat splenocytes to PHA and ConA was significantly lower than that of unchallenged conventional rats. Challenge with viable C. albicans did not result in a suppression of gnotobiotic rat splenocyte responses to PHA and ConA, but rather, the disseminated infection resulted in as much as fivefold increases in PHA or ConA-induced blastogenesis. These findings suggest that disseminated candidiasis is capable of suppressing blastogenesis in immunologically mature conventional rats and of improving lymphocyte blastogenesis from immunologically immature germfree rats.     

Mitogen- and antigen-specific proliferation of T cells in murine toxoplasmosis is inhibited by reactive nitrogen intermediates.

Acute and chronic infections with Toxoplasma gondii result in a nonspecific suppression of immunologic function in mice and humans. Proliferation of spleen cells in response to concanavalin A (ConA) and toxoplasma lysate antigen (TLA) was studied during the course of infection in mice susceptible (CBA/Ca) and resistant (BALB/c) to development of toxoplasmic encephalitis to determine if reactive nitrogen intermediates (RNI) are involved in the suppression of the proliferative responses. Maximal suppression of proliferation of spleen cells in response to ConA and TLA was observed on days 7 and 14 after infection and correlated with elevated levels of nitrite in spleen cell culture supernatants. By day 68 postinfection in BALB/c mice, proliferative responses returned to normal levels, whereas in CBA/Ca mice, they remained suppressed. The addition of an inhibitor of production of RNI (NG-monomethyl-L-arginine) increased proliferation of spleen cells in response to both ConA and TLA at days 7, 14, and 21 after infection. Depletion of adherent cells from spleen cell preparations obtained from acutely infected mice followed by their repletion with adherent spleen cells from uninfected mice resulted in increased proliferation of spleen cells from infected mice and a significant decrease in nitrite in the cultures. These results indicate that production of RNI by macrophages contributes significantly to the suppression of the spleen cell proliferation observed in the acute stage of toxoplasmosis.     

Acceleration of Immune Reconstitution after Bone Marrow Transplantation in Mice by Bone Marrow Stromal

To observe potential effect of the engineered bone marrow stromal cell line QXMSC1 secreting IL-6 (QXMSCIL-6) on accelerating immnune reconstitution in syngeneic bone marrow transplantation in mice, QXMSC1 was transfected with the eukaryocytic expression vector pcDNAIL-6, which contained hIL-6 cDNA by liposome-mediated gene transfecting technique. G418-resistance clone was selected by limiting dilution. The highest secreting clone was selected by ELISA assay and used in animal experiments. The recipient mice (BALB/c) were lethally irradiated and cotransplanted syngeneic bone marrow (10^7/mice) and the QXMSCIIL-6 (5&#215;10^5/mice). Lymphocyte proliferation induced by ConA and LPS, helper T lymphocyte precursor (HTLp), cytotoxic T lymphocyte precursor (CTLp), plaque-forming cell (PFC), delayed type hypersensitivity (DTH) were examined 30, 60 days in post transplantation respectively. The results showed that lymphocytes proliferation to ConA and LPS, HTLp, CTLp increased, DTH and PFC were improved by cografted stromal cells QXMSCIIL-6 on 30, 60 days after BMT. These results demonstrated that the bone marrow stromal cell line QXMSC1 IL-6 transfected with IL-6 (QXMSC11L-6) accelerated immnune reconstitution in syngeneic bone marrow transplantation.     

The stimulation of human B and T lymphocytes by various lectins

The response of human peripheral blood lymphocytes to different lectins was tested in vitro by monitoring DNA synthesis, blast transformation, and mitotic activity. One group of lectins - RCA, VGA, HPA, PNA, and UEA - showed no stimulating effects at all. WGA and VVA induced DNA synthesis and blast transformation but failed in stimulating mitosis. The mitogens PHA, ConA, LCA, and PWM showed peaks of mitotic activity at 50-60 hours for PHA, 70 hours for ConA, 80 hours for LCA, and between days 4 and 5 for PWM. The stimulation of different subpopulations of lymphocytes was investigated by immunological methods for the detection of B- and T-cell-specific surface structures during the whole incubation period. PHA proved to be a predominantly T cell stimulating agent, whereas ConA seemed to activate a higher proportion of B cells than yet known. PWM and the so-called T cell mitogen LCA turned up to stimulate a large number of B cells, but lead also to a T cell activation. The analysis of SCE events in stimulation experiments with these two lectins showed the early proliferation of a cell population with low SCE frequencies and the late propagation of a cell population with higher SCE rates. It could be assumed that the first population is represented by B- and the second by T-cells.     

芹菜素对小鼠T淋巴细胞体外活化和增殖的抑制作用

研究芹菜素(apigenin,AP)对小鼠T细胞体外活化和增殖的影响,探讨其作用机制,以及将其开发为免疫抑制药物的可能。无菌分离小鼠淋巴结细胞,加入不同浓度(25μmol/L、50μmol/L、100μmol/L、150μmol/L和200μmol/L)的芹菜素预孵育4 h,用多克隆刺激剂刀豆蛋白A(concanavalin A,ConA)诱导T细胞活化和增殖,并用MTT法检测该药物浓度对T细胞的毒性作用。荧光标记抗体双染色结合流式细胞术,检测各浓度AP对ConA诱导的小鼠T细胞表达早期、中期和晚期活化抗原CD69、CD25和CD71的影响;以二乙酰羧基荧光素-琥珀酰亚胺酯(carboxyfluoresceindiacetate-succinim-idyl ester,CFDA-SE)染色结合流式细胞术,检测各浓度AP对ConA诱导的小鼠T细胞增殖的影响,并应用ModFit软件分析其增殖指数(proliferation index,PI)。结果显示,25~200μmol/L AP对ConA诱导的T细胞CD69、CD25的表达有显著抑制作用(P<0.01),且呈剂量依赖关系;25μmol/L和50μmol/L的AP抑制CD71的表达,但高浓度(大于100μmol/L)才有显著抑制作用(P<0.01)。各浓度AP对ConA诱导的T细胞增殖均有显著抑制作用(P<0.01),且呈剂量依赖关系。表明在一定浓度范围内,AP可显著抑制ConA诱导的小鼠T细胞体外活化及增殖,有望通过进一步研究将其开发成免疫抑制药物。     

连翘提取物对小鼠T淋巴细胞体外活化与增殖的影响

目的:分析连翘(FS)提取物对小鼠淋巴结T细胞的体外活化与增殖的影响,初步探讨其免疫抑制作用机制.方法:无菌分离小鼠淋巴结细胞,加入多克隆刺激剂刀豆蛋白A(ConA)进行刺激,利用荧光标记的单克隆抗体(mAb)染色结合流式细胞术(FCM),检测小鼠T淋巴细胞的表达的活化抗原CD69、CD25、CD71的表达情况;以羧基荧光素乙酰乙酸琥珀酰亚胺酯(CFDA-SE)染色,以FCM分析FS对淋巴细胞体外增殖的影响.结果:终浓度为40、80、160 mg/L的FS均对ConA刺激诱导的T细胞CD69、CD25和CD71的表达有降低作用(P＜0.05).CFDA-SE染色分析显示,上述浓度的FS对ConA诱导的小鼠T淋巴细胞增殖具有抑制作用(P＜0.05).结论:FS对ConA诱导的T细胞早、中、后期活化和体外增殖有抑制作用.     

Systemic immunosuppression following a single pharyngeal aspiration of 1,2:5,6-dibenzanthracene in female B6C3F1 mice

1,2:5,6-Dibenzanthracene (DBA) is ubiquitous in our environment as a contaminant produced by incomplete combustion of organics from sources such as forest fires, cigarette smoke, and asphalt paving, and it is more immunosuppressive of the T-dependent antibody-forming cell (AFC) response than the well-studied polycyclic aromatic hydrocarbon, benzo(a)pyrene. The systemic immunosuppressive effects of DBA were investigated following a single pharyngeal aspiration (pa) in female B₆C₃F₁ mice. The immunotoxic effects of DBA were evaluated using numerous assays of varying complexity to evaluate innate (natural killer [NK] cell activity), cell-mediated (T-lymphocyte proliferation, mixed leukocyte response [MLR], cytotoxic T-lymphocyte [CTL] activity, delayed-type hypersensitivity [DTH]), and humoral immunity (B-lymphocyte proliferation, T-dependent antibody responses). A single pa of DBA at doses up to 30?mg/kg had no effect on NK cell activity, anti-CD3 antibody-mediated T-lymphocyte proliferation, the MLR, or B-lymphocyte proliferation. DBA at 30?mg/kg suppressed Concanavalin A (ConA)-stimulated T-lymphocyte proliferation and the CTL response. DBA exposure reduced cytokine production in spleen cell culture supernatants after in vitro stimulation with ConA or lipopolysaccharide (LPS). Immunosuppression was observed at lower doses in the holistic assays. The DTH response to Candida albicans was significantly decreased at 3.0?mg/?kg DBA, while the AFC response was intermittently suppressed at 1.0?mg/kg, with no effect observed at 0.3?mg/kg. These results demonstrate that a single pa of DBA produces systemic immunotoxicity, and of the assays utilized, the holistic assays (i.e., DTH, AFC) appear to be most sensitive to the immunosuppressive effects of DBA.     

Interleukin-15 production by monocyte-derived dendritic cells and T cell proliferation in HIV-infected patients with discordant response to highly active antiretroviral therapy

A discordant response to highly active antiretroviral therapy (HAART) occurs when CD4 T cell counts are stable or increased over time despite persistently detectable HIV-RNA levels. In order to identify immunological factors affecting discordant treatment responses, a total of 27 HIV-infected patients were studied: (a) 10 naive patients (mean CD4+ = 101.5 cells/microl; mean HIV-RNA = 4.8 log10 copies/ml); (b) seven responder patients (mean CD4+ = 908.9 cells/microl); and (c) 10 discordant patients (mean CD4+ = 396.1 cells/microl; mean HIV-RNA = 5.4 log10 copies/ml). Five healthy blood donors were included as HIV-seronegative controls. The following parameters were evaluated: interleukin (IL)-15 production by monocyte-derived dendritic cells (MDDC) after stimulation with lypopolysaccaride (LPS) and Candida albicans; recall and HIV-1-specific antigen lymphocyte proliferation (LP). Increased levels of IL-15 production by MDDC after stimulation with LPS and C. albicans were found both in discordant patients and responder patients. Conversely, a strong reduction of IL-15 levels was observed in naive patients. Discordant patients developed positive LP responses to C. albicans and HIV-1 p24. LP in response to C. albicans and HIV-1 p24 was also positive in responder patients. Decreased LP response was found in naive patients. In conclusion, HIV-infected patients with discordant viro-immunological responses to HAART present increased levels of IL-15 production by MDDC and enhanced recall and HIV-1-specific antigen LP responses, suggesting an improvement in indices of immune function.     

雌二醇对大鼠细胞免疫功能 的影响

本文采用大鼠去卵巢或再补充雌二醇(E_2)的方法探讨 E_(2)对免疫器官及其细胞免疫功能的影响。结果表明,去卵巢大鼠的脾脏和胸腺肥大,重量增加,脾细胞和胸腺细胞数明显增多。给去卵巢大鼠补充 E_2可使上述变化逆转。切除卵巢后的大鼠脾细胞对 ConA 诱导的增殖应答反应和诱生 IL—2的能力明显增高,补充 E_(2)亦可使上述变化逆转。但去卵巢大鼠的胸腺细胞对 ConA 诱导的增殖应答反应未能检测到可见的变化;给去卵巢鼠补充 E_2后,胸腺细胞对 ConA 的应答反应反而增强,并观察到胸腺细胞呈现轻度的自发增殖反应。实验结果表明:E_2能影响外周和中枢免疫器官及其细胞免疫功能,但这种影响在外周和中枢免疫器官表现出不同的变化。     

T cell hyperproliferation in autoimmunity prone obese strain (OS) chickens is independent of abnormal mitogen binding in vitro and can be demonstrated in vivo

In contrast to systemic autoimmunity, spontaneous autoimmune thyroiditis of Obese strain (OS) chickens is associated with a marked T cell hyperreactivity in vitro, i.e. an increased proliferation and interleukin 2 (IL 2) secretion in response to Concanavalin A (ConA). In the present study we report an enhanced capacity of OS peripheral lymphoid cells (splenocytes and peripheral blood lymphocytes, PBL) to adsorb fluorescein isothiocyante (FITC) labelled ConA, but not phytohemagglutinin (PHA). However, the elevated ConA binding cannot be a prerequisite for in vitro ConA hyperreactivity as OS thymocytes are normal with respect to ConA binding but nonetheless exhibit elevated responses to this mitogen. Moreover, ConA binding does not correlate with the frequency of cells able to express IL 2 receptors upon short term ConA stimulation. The percentage of ConA activatable cells was found to be increased in OS- PBL as compared to normal control PBL, but was unaltered in OS splenocytes. This finding points to a further mechanism of T cell hyperreactivity in OS chicks in addition to the previously reported defects in nonspecific immunosuppression. Finally, enumeration of cells in the S phase revealed that enhanced proliferation of OS T lymphocytes was not restricted to the in vitro response to ConA and phytohemagglutinin (PHA) but also occurs in vivo.