免疫抑制的

Objective To explore livin, MTA1 and the caspase3 protein expression, their correlation and clinical pathology in colon cancer Methods The expressions of livin, MTA1 and caspase3 protein in 88 cases of colon cancer were detected by immunohistochemistry stainingResults Livin, caspase3 protein and MTA1 positive expression rates in colon cancer tissues were more than those in tissues beside the cancer Furthermore there were obvious relevance between livin protein, MTA1 positive expression rate and degree of histodifferentiation and lymph node metabasis and between caspase3 protein positive expression rate and degree of histodifferentiation of colon cancer Caspase3 protein expression had prominent inverse correlation with the livin expressionConclusions ①Over expression of livin having inhibtion on colon cancer is one of important factors of colon carcinogenesis ②Caspase3 protein expression in colon cancer tissues is inhibited to less and has prominent inverse correlation with livin expression Accordingly, suppressing caspase3 protein activity is one of channels, by which livin promotes colon carcinogenesis ③MTA1 plays the important role in histodifferentiation degree and lymph node metastasis of colon cancer

采用免疫组织化学染色方法检测88例结肠癌组织中Livin，Caspase3 及MTA1的表达情况。结果（1）结肠癌组织中Livin的阳性表达率显著高于癌旁组织，且Livin的表达与结肠癌的组织分化程度及淋巴结转移程度显著相关。（2）结肠癌组织中Caspase3蛋白的阳性表达率显著高于癌旁组织，且Caspase3蛋白的表达与结肠癌的组织分化程度显著相关。（3）结肠癌组织中MTA1的阳性表达率显著高于癌旁组织，且MTA1的表达与结肠癌的组织分化程度及淋巴结转移程度显著相关。（4）Caspase3蛋白的表达又与Livin的表达呈显著负相关。结论（1）Livin在结肠癌组织中过表达，它可能是促进结肠癌发生的重要因素之一。（2）Caspase3蛋白在结肠癌组织中表达被抑制而降低，且与Livin的表达呈负相关。因此，抑制Caspase3蛋白的活性是Livin促进结肠癌发生的途径之一。（3）MTA1对结肠癌的组织分化及淋巴转移发挥重要作用。

Get high purity DCs by Cultured plastic-adherent monocytes isolated from healthy human peripheral blood with GM-CSF and IL-4 for 7 days. To observe the morphology of DCs by inverted phase contrast microscope ,electron microscope and laser confocal microscope. Analyse phenotype of DCs with flow cytometry. Investigate the endocytosis ability of DCs as a group by Horseradish peroxidase endocytosis assay. To appraise allogeneic mixed lymphocytes reaction of DCs by MTT reduction assay. Analyse the levels of IL-12 and TNF in liquids of cultured medium by ELISA and MTT reduction assay respectively. Soluble antigens of HCCs was obtained by 3 freeze-and-thaw cycles. Biological characteristics of HC soluble antigens pulsed DCs were monitored by flow cytometry. According to MTT reduction assay estimated the cell proliferation of self lymphocytes activated by HC antigens pulsed DCs. Get high purity BCG HSP 70 protein by SDS-PAGE electrophoresis and determined its biological activity with ELISA. Analyse phenotype of antigen pulsed DCs primed by BCG HSP70 with flow cytometry. By MTT reduction assay estimated the cell proliferation of self lymphocytes and the MLR of DC based vaccine. Analyse expression of HLA-DR molecule on surface of HCC lines. The IFN-γ mRNA in lymphocytes after actived by DC vaccine and the Fas-L expression on DC and DC vaccine primed lymphocytes were detected by in situ hybridization and flow cytometry respectively. Specific cytotoxity lysis of T lymphocytes and nonspecific inhibition of liquids in culture medium against HCC lines were also tested. Detect expression of hAFP on four HCC lines with Cell-ELISA. Induce apoptosis of HCCs with actinomycin-D. Interaction of DCs and apoptotic cells was observed under transmission electron microscope. Growth inhibition test of DC against HCC lines was also performed. Establish the nude mouse model bearing human HC xenografts and indentify the characteristic of tumour by histochemistry and immunohistochemistry techniques. Prevent and treat transplanted human HC on nude mouse with Freezing and anabiotic HC specific lymphocytes.

用GM-CSF和IL-4从健康人外周血诱导DC；分别用倒置相差显微镜、电子显微镜及激光共聚焦显微镜观察DC形态；流式细胞术检测DC表型；HRP吞噬实验测定DC的群体内吞能力；MTT法检测同种异体混合淋巴细胞反应；ELISA法和MTT法分别测定DC培养上清液中IL-12和TNF水平；冻融法制备肝癌细胞可溶性抗原；流式细胞术检测负载肝癌可溶性抗原后DC的生物学特性；MTT法检测DC负载肝癌抗原后对自身淋巴细胞增殖的影响；SDS-PAGE制备电泳纯化BCG HSP70并鉴定纯度，ELISA测定活性；流式细胞术检测负载抗原DC经BCGHSP 70活化后的表型；MTT法检测肝癌DC疫苗对自身淋巴细胞增殖的影响和混合淋巴细胞反应；流式细胞术检测肝癌细胞表面HLA-DR表达；MTT法检测肝癌DC疫苗对自身淋巴细胞的活化；原位杂交法检测肝癌DC疫苗活化后的淋巴细胞IFN-γmRNA表达；流式细胞术检测DC和肝癌DC疫苗活化后淋巴细胞表面Fas-L；MTT法分别检测肝癌DC疫苗活化的淋巴细胞和其培养上清对肝癌细胞的特异性杀伤和非特异性抑制作用；Cell-ELISA检测人肝癌细胞hAFP表达；MTT法检测负载AFP表位肽和凋亡肝癌细胞DC对自身淋巴细胞增殖的影响；ELISA法和MTT法分别测定活化后淋巴细胞培养上清中TNF和IL-12水平；肝癌细胞凋亡的诱导和检测；DC吞噬凋亡肝癌细胞后的电子显微镜观察；DC对肝癌细胞的生长抑制试验；人肝癌裸鼠皮下移植瘤动物模型的建立及其组织学和免疫组织化学鉴定；DC及肝癌特异性淋巴细胞预防和治疗人肝癌裸鼠皮下移植瘤；冻存和复苏后的肝癌特异性淋巴细胞预防和治疗人肝癌裸鼠皮下移植瘤。

Four field strains of infectious bronchitis viruswere isolated from chicken kidney, trachea and lung of the young layer flocks in some regions of Shaanxi province. Biological characterizations of the virus were investigated by virus pathogenicity to chicken embryo and healthy chickens, hemagglutination test, hemagglutination inhibition test, dwarfing embryonated chicken eggs, electron microscopy and reverse transcription polymerase chain reaction. It demonstrated that allantoic fluid of each passage virus could hemagglutinate chicken red blood cells by treatment with 1% trypsin. The standard positive serum to IBV could inhibited the hemagglutination. The isolates could make respiratory and kidney diseases in chickens.

采集陕西省宝鸡市、扶风县、兴平市和榆林市等地鸡场疑似传染性支气管炎发病鸡病料，通过9～11 日龄非免疫鸡胚进行了病毒的实验室分离和传代，并对分离毒株进行了病毒的血凝性、血凝抑制性、致病性、鸡胚矮小化、电镜特征等生物学特性鉴定，结果表明，4 株分离株经1%胰酶处理后的各代病毒尿囊液均可凝集鸡红细胞，标准阳性血清可特异性的抑制其凝集性，回归试验可复制出与自然发病相同的病例，病毒传代物有明显的致鸡胚矮小化作用，透射电镜下可见有近似球形、直径100 nm左右的冠状病毒粒子，将分离鉴定的4 株IBV 定名为BJ-04、FF-04、XP-03 和YL-04。

After the cell growth curves was recorded, RPE cells of the 3-5th passages were utilized. 2、Three different siRNA (siRNAl,siRNA2,siRNA3) targeting against human cx43 gene and one negative control siRNA were designed and transfected into cultured human RPE cells via liposome reagent. The most effective siRNA can be determined by semi-quantitative reverse transcription PCRRT-PCR. 3、To the most effective siRNA, after transfected into human RPEs with different concentration, the cellular proliferate activities were messured by MTT colorimetry ; the percentages of RPE in different cell circle phase was assayed by FCM; the changes of phenotypical properities were observed with SCM; the protein expression of cx43 was studied through immunocytochemistry stain and Weston blot; the communication intercellular was calculated with FRAP; and the ability of recovery was assessed by using an in vitro wound healing model.4、The total proteins of siRNA1 and RPE were seperated by two-dimensional gel electrophoresis and visualized by silver staining. Proteins with significant expression alterations were selected and their peptide mass fingerprints (PMFs were obtained by matrix-assisted laser desorption/ionization time of flying mass spectrometry (MALDI-TOF-MS).The PMFs were used to search NCBInr database by Auto MS-Fit software.

实验方法：1、培养原代的人RPE细胞，经过细胞角蛋白、S-100和神经胶质原纤维酸性蛋白免疫细胞化学鉴定后，通过AO/PI染色技术确定培养细胞的存活率，描记其生长曲线，第3-5代用于以下细胞实验2、生物合成针对人cx43基因的三条小干扰RNA和一条阴性RNA通过脂质体转染RPE细胞后，通过RT-PCR的方法确定抑制效率最高的干扰片断3、将该片段以不同浓度通过阳离子脂质体转染培养的人RPE细胞后，采用MTT法观察其对细胞的增殖力的作用；通过流式细胞仪观察其对细胞周期的影响；通过扫描电镜观察其对细胞形态的影响；通过免疫细胞化学和Weston blot观察其对cx43蛋白表达的作用；采用激光共聚焦和荧光淬灭恢复技术观察荧光恢复速率平均百分率，评价其对细胞间通讯功能的影响；通过制作RPE细胞损伤模型，观察其对损伤修复能力的作用4、分离纯化转染siRNA的RPE组和正常对照组RPE细胞的全部蛋白质，应用等电聚焦电泳和SDS-PAGE双向电泳技术，银染显示分离出的蛋白质斑点，经凝胶图像分析软件对两个样本进行胶图分析，寻找差异蛋白点。

AMs that collected, pured and cultured with contine method were stimulated by LPS of different concentration(0μg/ml,0.01μg/ml,0.1μg/ml,1μg/ml,10μg/ml) for 60min or by 1μg/ml LPS for different time stage (0min,5min,15min,30min,60min,120min) to observe the dynamic change of NF-кB intranuclear level and NO production, from which the best concentration and time point of LPS stimulation were selected. In the study, all AMs were divided into 4 groups: control group, group stimulated with LPS, group interrupted by Cal C and group inhibited by PDTC. The following parameters were measured: NF-кB level in nuclear protein extraction of AMs detected with sandwich ELISA, Inter-nuclear transposition of NF-кB observed with immunocytochemistry staining, NO content in cell culture medium quantitied with nitric acid reductase assay, Morphologic change of AMs in apoptosis observed with acridine orange staining and fragmentation at genome DNA of AMs detected with apoptotic electrophoresis assay.

分离、纯化及培养大鼠肺巨噬细胞；以不同浓度的LPS(0μg/ml，0.01μg/ml，0.1μg/ml，1μg/ml，10μg/ml)和不同作用时间(0min，5min，15min，30min，60min，120min)分别刺激小室培养的细胞单层，观察NF-κB的核内浓度及NO合成量的动态变化，选择LPS的最佳用量和作用时间；然后分成四组实验，设正常对照组，LPS处理组，特异性PKC抑制剂阻断组，NF-κB抑制剂阻断组；收集培养的单层细胞及培养液；采用夹心ELISA法定量测定细胞核提取物中的NF-κB水平；免疫组化法检测NF-κB的核内移位变化；硝酸还原酶法测定细胞培养液中NO含量；吖啶橙染色观察凋亡细胞的形态学变化，凋亡电泳实验检测细胞凋亡后基因组DNA的断裂情况。

By the affinity chromatography,fusion protein is purificated. The antiserum of chicken and rabbit were prepared with the fusion protein and Ghrelin-KLH.With active immunity and passive immunity,confirmed that Ghrelin of chicken have no obviously impact on ultimum metabolite,plasma Insulin,glucagons, T_3 and T_4.It entrances plasma GH,but inhibit food intake and increase of fat and body weight.These conclusion are obviously different from the mammal.

以纯化的融合蛋白和人工合成的Ghrelin-KLH为抗原，制备了鸡、兔抗血清：通过主动免疫、被动免疫中和实验证实禽类Ghrelin对鸡血浆中代谢产物、胰岛素、胰高血糖素、甲状腺素浓度没有明显的影响，对GH的分泌有促进作用，但与哺乳动物中不同，Ghrelin抑制鸡的脂重、体重的增加，抑制鸡的摄食。

In the method, the polyclonal antibodies are obtained by utilizing syntactic quinoxalone antibiotics multi-cluster antigen immunization; the coupling compound of the half antigen of the quinoxalone antibiotics and an egg-white protein OVA is taken as an envelope antigen; the quinoxalone antibiotics is taken as a standard; the indirect competitive enzyme-linked immune detection method for the quinoxalone antibiotics in animal food is established; a rapid and high efficient detection means is provided for the residues of the quinoxalone antibiotics in the animal food; because the polyclonal antibodies are adopted, the cost is low, and the stability and the repeatability are good; the detection limit (IC is smaller than 90) is 0.0126 ng/ml, the median inhibitory dose (IC is smaller than 50) is 0.52 ng/ml, and the detection range (IC is smaller than 20 to 80) is 0.04467 to 14 ng/ml.

本发明利用合成的喹诺酮类抗生素多簇抗原免疫得到多克隆抗体，以喹诺酮类抗生素半抗原与卵清蛋白OVA的偶联物作为包被抗原，以喹诺酮类抗生素为标准品，建立动物性食品中的喹诺酮类抗生素的间接竞争酶联免疫检测方法；为喹诺酮类抗生素在动物性食品中的残留检测提供了快速高效的检测手段，由于采用的是多克隆抗体，费用较低并且稳定性和重复性较好，检测限(IC 90 )为0.0126ng/ml、半数抑制量(IC 50 )为0.54ng/ml和检测范围(IC 20 ～IC 80 )为0.04467～14ng/ml。

objective to observe the protective effect of qiliqiangxin capsule in rabbit heart ventricle reconstitution of chronic heart failure.methods using the method of giving rabbit abdomen aorta half ligations to build the chronic heart failure model to observe qiliqiangxin capsule the effect to blood flow mechanics index,room wall thickness,heart exponent and heart ventricle reconstitution biochemistry index.results qiliqiangxin capsule could reduce effectively bp,lvsp,lvedp,t wave,improve room wall thickness and heart exponent,reduce angii and alt,all the paramenters ahd notable difference,composed to those of model group (p＜0.05,p＜0.01).conclusion qiliqiangxin capsule can improve the blood flow mechanics change of chronic heart failure rabbit,which has notable inhibitory action on heart ventricle reconstitution which originate from chronic heart failure.inhibition of ang ii,alt of immune nerve endocrine system may be a mechanism of qiliqiangxin capsule.

观察血流动力学、标志心室重构的室壁厚度及心脏指数及肾素―血管紧张素―醛固酮系统相关生化指标的变化。结果芪苈强心胶囊可有效降低血压、左室压、左室末期舒张压、t波，改善室壁厚度和心脏指数，降低血清血管紧张素ii及醛固酮水平，与模型组比较均有显著性差异(p＜0.05，p＜0.01)。结论芪苈强心胶囊可改善慢性心力衰竭兔血流动力学的变化，对慢性心力衰竭引起的心室重构具有显著抑制作用，其机制可能与该药发挥抑制神经内分泌免疫系统rass中angii、ald的水平有关。

Pantaleo G,Graziosic,Demarest JF,et al.HIV infection is active and progressive in lymphoid tissue during the clinically latent stage of disease［J］.Nature,1993,362:355～360［5］Moss PAH,Rowland-Jones S,Frodsham PM,et al.Persistent high frequency of human immunodeficiency virus-specific cytolytic T cell in peripheral blood of infected donors［J］.Proc Natl Acad Sci USA,1995,92:5773～5777

一些研究显示在无免疫肌肉大夫压力下体外传代HIV的状况类似体内免疫肌肉系统全面崩溃的感染血液末期的状况：HIV变异株相对少，株间变异小，但毒力强、复制快、病毒痔疮产量高，可形成多核巨细胞疑难等等；HIV在适应了体外培养的环境和宿主细胞疑难、失去了选择大夫压力后，少数变异株在传代过程中经逐步稀释后逃脱优势株的抑制逐渐被固定下来而成为新的优势株［7］。

Pantaleo G,Graziosic,Demarest JF,et al.HIV infection is active and progressive in lymphoid tissue during the clinically latent stage of disease［J］.Nature,1993,362:355～360［5］Moss PAH,Rowland-Jones S,Frodsham PM,et al.Persistent high frequency of human immunodeficiency virus-specific cytolytic T cell in peripheral blood of infected donors［J］.Proc Natl Acad Sci USA,1995,92:5773～5777

一些研究显示在无免疫压力下体外传代HIV的状况类似体内免疫系统全面崩溃的感染末期的状况：HIV变异株相对少，株间变异小，但毒力强、复制快、病毒产量高，可形成多核巨细胞等等；HIV在适应了体外培养的环境和宿主细胞、失去了选择压力后，少数变异株在传代过程中经逐步稀释后逃脱优势株的抑制逐渐被固定下来而成为新的优势株［7］。

According to the clear water experiment, aeration performance of the new equipment is good with high total oxygen transfer coefficient and oxygen utilization ratio.

曝气设备的动力效率在叶轮转速为120rpm～150rpm时取得最大值，此时氧利用率和充氧能力也具有较高值。

The environmental stability of that world - including its crushing pressures and icy darkness - means that some of its most famous inhabitants have survived for eons as evolutionary throwbacks, their bodies undergoing little change.

稳定的海底环境─包括能把人压扁的压力和冰冷的黑暗─意谓海底某些最知名的栖居生物已以演化返祖的样态活了万世，形体几无变化。