滤泡辅助性T细胞在特发性血小板减少性紫癜患者的变化及其意义

目的：研究滤泡辅助性T细胞（T follicular helper cells,Tfh）在免疫性血小板减少性紫癜（immune thrombocytopenic purpura,ITP）患者的表达并探讨其临床意义。方法：用流式细胞术检测20例健康人、25例ITP患者外周血CXCR5＋CD4＋T细胞占CD4＋T细胞的比例。结果：与健康对照组相比,ITP患者外周血CXCR5＋CD4＋T细胞占CD4＋T细胞的比例显著增高（P〈0.05）。结论：Tfh在ITP患者外周血比例增高,为Tfh能否为ITP的免疫调节和干预提出新的方向提供了证据。     

滤泡辅助性T 细胞在特发性血小板减少性紫癜患者的变化及其意义*

目的:研究滤泡辅助性T细胞(T follicular helper cells,Tfh)在免疫性血小板减少性紫癜(immune thrombocytopenic purpura,ITP)患者的表达并探讨其临床意义。方法:用流式细胞术检测20例健康人、25例ITP患者外周血CXCR5+CD4+T细胞占CD4+T细胞的比例。结果:与健康对照组相比,ITP患者外周血CXCR5+CD4+T细胞占CD4+T细胞的比例显著增高(P<0.05)。结论:Tfh在ITP患者外周血比例增高,为Tfh能否为ITP的免疫调节和干预提出新的方向提供了证据。     

Characterization of a subclone (D10S) of the D10.G4.1 helper t-cell line which proliferates to attomolar concentrations of interleukin-1 in the absence of mitogens

Most studies have shown that interleukin-1 (IL-1) acts as a helper or co-stimulator in T-lymphocyte activation and proliferation by mitogens or antigens. We describe here a stable subclone (D10S) of the murine D10.G4.1 helper T-cell which proliferates to subfemtomolar (attomolar) concentrations of IL-1 beta or alpha in the absence of mitogens. D10S cells have been maintained in culture for over two years without splenic cell feeder layers nor antigen stimulation. Detection of proliferation can be made by either uptake of tritiated thymidine at 72 h or in 48 h by a colorimetric assay which measures mitochondrial dehydrogenases; the latter assay is rapid and inexpensive. D10S cells are distinct from the parent clone D10.G4., which requires mitogens for IL-1 activity. IL-1-induced proliferation is independent of the elaboration of IL-2, IL-4, or IL-6, although these cells proliferate to these lymphokines at considerably higher concentrations when compared to IL-1. The D10S cells proliferate in direct correlation to the duration of IL-1 presence in the culture. We found no evidence that IL-1 induced more IL-1 in these cells. The subclone is highly specific for IL-1: proliferation was not observed to endotoxin, human or murine interferon-gamma (IFN gamma), tumor necrosis factor (TNF), lymphotoxin, or granulocyte-macrophage colony stimulating factor (GM-CSF). There was no suppressive effect of transforming growth factor (TGF beta). Only at high concentrations (100 ng/ml) did IL-6 induce proliferation. We conclude that this stable, feeder layer-free cell line is highly sensitive to IL-1 which acts as a direct stimulant for these cells; they are also useful for bioassays as well as the study of IL-1 receptors as described in the accompanying paper.     

IL-6-induced Bcl6 variant 2 supports IL-6-dependent myeloma cell proliferation and survival through STAT3

IL-6 is a growth and survival factor for myeloma cells, although the mechanism by which it induces myeloma cell proliferation through gene expression is largely unknown. Microarray analysis showed that some B-cell lymphoma-associated oncogenes such as Bcl6, which is absent in normal plasma cells, were upregulated by IL-6 in IL-6-dependent myeloma cell lines. We found that Bcl6 variant 2 was upregulated by STAT3. ChIP assay and EMSA showed that STAT3 bound to the upstream region of variant 2 DNA. Expression of p53, a direct target gene of Bcl6, was downregulated in the IL-6-stimulated cells, and this process was impaired by an HDAC inhibitor. Bcl6 was knocked down by introducing small hairpin RNA, resulting in decreased proliferation and increased sensitivity to a DNA damaging agent. Thus, STAT3-inducible Bcl6 variant 2 appears to generate an important IL-6 signal that supports proliferation and survival of IL-6-dependent myeloma cells.     

Immune characteristics study of AG490, a signal pathway inhibitor,in EAE model mice

Multiple sclerosis (MS) is an autoimmune disease characterized by demyelination, axonal damage and progressive neurologic dysfunction in central nervous system (CNS). Many evidences show that B cells play an important role in the pathogenesis of MS. Follicular helper T cells (Tfh) secrete IL-21 to prompt the proliferation and differentiation of B cells in germinal center (GC) through clonal proliferation, somatic hypermutation, antibody class switching, antibody affinity maturation process. AG490 is a synthetic inhibitor to JAK-STAT signal pathway, which has been studied in inflammatory, tumor and autoimmune diseases. In the present study, the experimental mice were divided into 3 groups, vehicle group and AG490 group were given MOG35-55 to induce EAE model, from the third day after immunization, the mice were given vehicle or AG490 by intraperitoneal injection every other day. All mice were assessed clinical scores after immunization. On twentieth day, all mice were sacrificed, HE staining and solochrome cyanine staining were performed to evaluate inflammatory cells infiltration and demyelination, spleen sections were stained with PNA-FITC to analyze the difference in germinal center. Compared with vehicle group, the incidence of AG490 group was deceased, onset time was delayed, the severity was significantly reduced. The inflammatory cells and demyelination in AG490 group were lower than those in vehicle group. Immunofluorescence showed the fluorescence intensity of AG490 group was significantly lower than in the vehicle group, but higher than that of control group.     

IL-8与胃癌关系的研究进展

IL-8是趋化性细胞因子,可以促进炎症细胞趋化和诱导细胞增殖,在胃癌组织中高表达。影响IL-8在胃癌组织中表达水平的因素包括幽门螺杆菌感染、NF-κB的干预、IL-8的基因多态性、表皮生长因子（EGF）的水平。IL-8主要参与胃癌的血管形成、促进金属蛋白酶的表达、影响E-cad和ICAM-1的水平、促进细胞有丝分裂、促使肿瘤转移等,对胃癌的发生、发展、预后均起重要的作用。选择性阻断IL-8及其受体具有抑癌作用,有望成为胃癌治疗的基础药物之一。     

趋化因子共受体在1型人免疫缺陷病毒感染中的作用

1型人免疫缺陷病毒(HIV-1)感染靶细胞是一个包含病毒膜蛋白和细胞膜受体相互作用的多极化过程,CCR5和CXCR4作为趋化因子受体参与这一过程,并且是M嗜性和T嗜性HIV-1感染的重要共受体。文章总结了作者在HIV-1共受体方面的工作,对趋化因子受体作为新的治疗HIV-1感染的工具的最新进展做了简要综述。     

The chemokine,CXCL16, and its receptor,CXCR6, are constitutively expressed in human annulus fibrosus and expression of CXCL16 is up-regulated by exposure to IL-1ß in vitro

Chemokines are an important group of soluble molecules with specialized functions in inflammation. The roles of many specialized chemokines and their receptors remain poorly understood in the human intervertebral disc. We investigated CXCL16 and its receptor, CXCR6, to determine their immunolocalization in disc tissue and their presence following exposure of cultured human annulus fibrosus cells to proinflammatory cytokines. CXCL16 is a marker for inflammation; it also can induce hypoxia-inducible factor 1α (HIF-1α), which is a phenotypic marker of heathy nucleus pulposus tissue. We found CXCL16 and CXCR6 immunostaining in many cells of the annulus portion of the disc. Molecular studies showed that annulus fibrosus cells exposed to IL-1ß, but not TNF-α, exhibited significant up-regulation of CXCL16 expression vs. control cells. There was no significant difference in the percentage of annulus cells that exhibited immunolocalization of CXCL16 in grade I/II, grade III or grade IV/V specimens. The presence of CXCL16 and its receptor, CXCR6, in the annulus in vivo suggests the need for future research concerning the role of this chemokine in proinflammatory functions, HIF-1α expression and disc vascularization.     

IL-18基因转染对大鼠C6胶质瘤细胞生长特性的影响

探讨IL-18基因转染对大鼠C6胶质瘤细胞生长特性的影响。用MTT法和流式细胞术检测C6／IL-18细胞和C6细胞的增殖特性和细胞周期分布。免疫细胞化学检测C6／IL-18细胞和C6细胞的增殖细胞核抗原(PCNA)、波形蛋白表达。结果显示,与C6细胞相比C6／IL-18细胞的增殖能力降低,G0/G1期细胞增多而G2／M期细胞减少;PCNA、波形蛋白表达降低。研究表明,IL-18基因具有抑制C6胶质瘤细胞增殖、降低其恶性程度的作用。     

人IL-6基因的克隆及其在原核生物中表达及条件的优化

构建人白细胞介素6(IL-6)的原核表达载体并优化其表达条件,为IL-6的高效表达提供试验依据。以人T细胞cDNA为模板通过PCR方法扩增IL-6基因,将其克隆到原核表达载体pET28a(+)中,酶切及测序鉴定重组体。将构建好的重组质粒转化大肠杆菌BL21(DE3),用IPTG进行诱导表达,产物用Western blotting及人IL-6检测试剂盒分析鉴定。在保持菌种不改变的前提下,分别改变IPTG的浓度、培养时间、卡那霉素浓度、培养温度等来优化IL-6表达条件。结果显示,原核表达载体pET28 a(+)-IL-6成功构建,可在大肠杆菌BL21(DE3)中诱导表达,得到相对分子质量约22 kD的IL-6蛋白,经Western blotting鉴定正确,经人IL-6试剂盒检测显示具有较高的免疫活性。在IPTG浓度400μmol/mL,卡那霉素浓度50μg/mL,40℃培养6 h的条件下,目的蛋白表达量最高,可占总蛋白表达量的40%。成功构建人IL-6原核表达载体且获高效表达,为研究IL-6生物学活性及产品开发提供了试验基础。     

蛇毒与细胞因子研究进展

全世界共有蛇类2500余种,其中毒蛇约650余种,估计每年被毒蛇咬伤的人数在30万以上,死亡率约为10％。我国蛇类有160余种,其中毒蛇约有50余种。剧毒、危害剧大的有10种,如眼镜蛇王、金环蛇、眼镜蛇、五步蛇、银环蛇、蝰蛇、蝮蛇、竹叶青、烙铁头、海蛇等,咬伤后能致人于死亡。我国两广地区蛇害严重,每年蛇咬伤的发病率约为25/10000。蛇毒的成分比较复杂．主要由蛋白质、多肽类和多种酶类组成。蛇毒对机体的作用比较复杂,按其有毒成分的毒理作用可分为神经毒、血循环毒和混合毒三类。银环蛇、金环蛇、海蛇的蛇毒主要含神经毒,其主要作用特点为通过多种不同的方式阻断神经一肌肉接头的冲动传递而导致呼吸肌麻痹,是蛇伤致死的主要原因;蝰蛇、五步蛇、烙铁头和竹叶青的蛇毒主要含血循环毒,包括心脏毒、凝血毒、溶血毒、蛋白水解酶、透明质酸酶等;眼镜王蛇等蛇毒属于混合毒,此类蛇毒既含神经毒成分,又含血循环毒成分。     

白细胞介素-6对新生大鼠海马神经元电压依赖离子通道和NMDA电流的影响

目的: 研究白细胞介素-6对海马神经元电压依赖离子通道和NMDA电流的影响.方法: 应用全细胞膜片钳技术观察IL-6对电压依赖性钠通道电流(INa),延迟整流性钾通道电流(IK),电压依赖性钙通道电流(ICa),NMDA(N-methyl-D-aspartate)受体通道电流的影响.结果: 50 ng/ml IL-6作用24 h后IK 和ICa明显减小,Cm明显增大.50,500 ng/ml时减小NMDA电流.结论: IL-6通过作用于电压依赖钾通道,钙离子通道及NMDA通道影响神经元功能.     

中药芪龙风湿丸对类风湿关节炎患者外周血细胞因子IL-1,IL-6,TNF-α的影响

目的采用中药为主的方法治疗类风湿性关节炎(RA),并进行患者外周血细胞因子(CK)的测定及观察治疗前后的变化。方法随机选择RA病人62例,其中治疗组42例以中药益气养阴,甘润辛通法为主自拟中药芪龙风湿丸治疗,同时予MTX每周5 mg,连服60天;对照组20例,予MTX每周5 mg,连续服用60天。同时观察药物对患者外周血细胞因子IL-1,IL-6,TNF-α的影响。结果两组外周血细胞因子治疗前均有异常表达,治疗后异常水平均有下降,中药组治疗前后自身比较,三种细胞因子的下调均有非常显著意义(P<0.01)。与对照组比较,治疗后中药组IL-1,TNF-α的下降有显著意义(P<0.05)。结论以益气养阴,甘润辛通法治疗RA,能改善外周血异常细胞因子表达,对RA病人临床症状和实验室指标均有一定改善作用。     

奶牛IL-6基因克隆与原核表达

目的:克隆新疆地区奶牛il-6基因.获得原核表达IL-6蛋白.方法:根据GenBank上牛IL-6的序列,用premier 5.0软件设计一对引物.以牛肺巨噬细胞提取的RNA反转录产物(cDNA)为模板扩增出567bp的条带,克隆到pBS-T载体,测序正确后,提取质粒经BamH Ⅰ和EcoR Ⅰ双酶切回收目的条带,亚克隆到pGEX-4T-1原核表达载体,转化DE3菌,经IPTG诱导表达.以表达产物免疫小白鼠血清为一抗榆测表达产物的反应原性.结果:成功克隆了新疆地区奶牛il-6基因,克隆部分编码188个氨基酸与GenBank公布的序列100%同源,表达出46 kDa的融合蛋白,免疫印迹检测显示原核表达牛IL-6有免疫原性.结论:新疆地区奶牛在IL-6的基因序列上与其他地方牛无差异,原核表达奶牛IL-6蛋白为在奶牛乳房炎疫苗中的佐剂效果研究打下了基础.     

离心机训练对大鼠脑及其它组织IL-6和TNFα基因表达水平的影响

为了解离心机训练前后大鼠脑及心、肺、肾和小肠组织中IL 6和TNFα基因表达水平的变化 ,对雄性SD大鼠进行动物离心机训练 ,刺激值从 + 7Gz～ + 12Gz ,每天增加 + 0 5Gz ,第 12d重复 + 12Gz刺激 ,第 13d离心机训练后 ,断头处死 ,取心、脑、肺、肾和小肠组织 ,分别提取mRNA并定量 .用地高辛标记IL 6和TNFαcDNA作为探针 ,进行狭缝杂交 ,杂交结果通过光密度扫描定量后 ,进行统计学处理 .离心机训练不同时间 ,比较大鼠各组织IL 6和TNFα基因表达水平的变化 .结果显示随着训练时间的延长 ,表达水平均有所差异 .提示训练对大鼠脑及其它主要组织IL 6和TNFα基因表达水平有影响 ,这种影响可能与机体对加速度作用的习服有关     

应用椭偏光学显微成像对IL-6与其受体的相互应用的初步观察

白细胞介素 6 (ＩＬ 6 )是一种具有复杂生物功能的细胞因子 ,可由多种淋巴类和非淋巴类细胞产生。它对机体多种组织及细胞均有不同程度的作用[1～ 3 ] 。近年来发现 ,临床上免疫异常性疾病 ,如发热、淋巴结肿大、血沉增快、急性期蛋白增高、高γ球蛋白血症、自身抗体阳性等症状都与ＩＬ 6的异常表达密切相关。ＩＬ 6的生物活性是通过细胞膜表面特异性受体介导的[4] 。研究ＩＬ 6与其受体的相互作用对于揭示某些疾病的发病机制 ,监测疾病进程以及指导临床治疗等均具有重要意义。用于研究ＩＬ 6与其受体相互作用的方法主要有ＩＬ 6依赖株细胞…     

用一个简单快速的RT-PCR技术筛选白细胞介素-6作用相关基因

为了研究白细胞介素-6(IL-6)作用相关基因以及一些可能受IL-6调控的基因,利用一个简单快速的以PCR为基础的方案,检测了IL-6处理和未处理的Sko007细胞中基因表达的差异,克隆并鉴定了差异表达基因的cDNA片段.首先用6-mer寡核苷酸引物进行反转录从而最大限度地将mRNA编码区序列生成cDNA;然后用2或3个较长的随机引物进行PCR扩增,并以不同引物组合重复PCR增扩;扩增产物在2%琼脂糖凝胶上电泳分离,回收差异片段并直接用于克隆、测序及进一步分析.在此研究中,获得了3个表达序列标签(EST),其中一个为新的基因片段,反向RNA杂交有力证实了它们与IL-6作用的相关性.进一步的生物信息学分析表明,新基因片段STRF17在多种组织中表达.