制备性分段电泳纯化基因工程白细胞介素—2

采用具有高分辨率特性的聚丙烯酰胺凝胶为载体,以电泳技术将工程菌株的各种蛋白成分精细分离,再用紫外检测手段对白细胞介素—2(IL—2)基因表达产物在凝胶上定位,通过准确地切割凝胶,提取其中的蛋白质,获得了高纯度(>95%)的基因工程IL—2产物.经用同位素掺入法测定,其对小鼠胸腺细胞有明显的促进增殖活性     

IL-2在自然感染兔脑炎原虫獭兔体内免疫作用的研究

为观察 IL-2在自然感染兔脑炎原虫獭兔体内的免疫作用 ,用双抗体夹心 EL ISA试剂盒对 8只自然感染兔脑炎原虫的獭兔和 4只健康长耳白对照兔的血清及培养 3 d和 4d的脾和淋巴结的细胞培养上清中的 IL-2进行了检测。检测结果为病兔与对照兔 IL -2平均含量差异不显著 (P >0 .0 5) ,但病兔 IL -2平均含量高于对照兔。提示 IL -2对病兔产生了一定的免疫作用 ,但可能不扮演主要角色     

白细胞介素-1β蛋白在孕哺期铅暴露仔鼠海马组织中的表达

为探讨母体铅染毒对其子一代海马组织中白细胞介素-1β（IL-1β）蛋白表达的影响,将雌性小鼠自妊娠第1天开始经饮水染铅（0.3,1.0,3.0g/L,对照组饮蒸馏水）至仔鼠出生后21d断乳为止。随机抽取各组仔鼠,于出生后7,14,21d分别测其血液和海马组织中铅的含量;于出生后21d,采用Western blot方法测定各组海马组织中IL-1β的表达情况。结果表明,孕哺期不同剂量铅暴露后,出生后7,14,21d的仔鼠血铅、海马铅水平均明显高于对照组（P〈0.05）。Western blot结果显示,IL-1β蛋白在不同剂量铅暴露组海马组织中的表达明显高于对照组（P〈0.05）。相关分析表明,IL-1β蛋白的表达与血铅、海马铅呈显著正相关（P〈0.05）。母体铅暴露使铅在仔鼠体内蓄积,提示仔鼠血铅和海马铅的升高,引起海马组织中IL-1β表达的上调,可能损伤了海马神经元,进而损伤了神经系统。     

Enhanced IL-6 transcriptional response to adenosine receptor ligands in horses with lower airway inflammation

Reasons for performing study: Accumulation of extracellular adenosine has been closely associated with human asthmatic responses. However, the relevance of adenosine signalling in equine airways has not previously been investigated. Objectives: To determine the expression of adenosine receptors (AR) in bronchoalveolar lavage (BAL) cells and assess the reactivity of these cells to AR ligands ex vivo, employing IL‐6 as readout of adenosinergic inflammatory signalling. Methods: Eight horses with varying degrees of lower airway inflammation and 10 healthy controls were analysed. Expression of AR‐subtypes in each BAL sample was determined by quantitative RT‐PCR and compared to that in 13 other tissues. Bronchoalveolar lavage cells were stimulated either with the adenosine analogue NECA, CGS‐21680 (A_2AAR selective agonist) or with a combination of NECA and SCH‐58261 (A_2AAR antagonist) and IL‐6 expression assessed. Results: Bronchoalveolar lavage cells predominantly expressed A_2BAR, with lower A_2AAR levels and marginal A₃AR expression; A₁AR was not detected. This pattern was similar to that of PBMCs but different from the other tissues tested. No significant differences in AR expression in BAL cells from both groups were detected, although a trend for decreased A_2BAR in airway‐compromised horses was observed. Treatment of BAL cells with the nonselective agonist NECA upregulated IL‐6 expression in cells from airway‐compromised horses, but levels remained unchanged in control animals. Furthermore, blockage of A_2AAR with SCH‐58261 enhanced IL‐6 mRNA induction by NECA in both groups, with higher levels in airway‐compromised horses; the amplitude of this response correlated with neutrophil count. Conclusions: These results demonstrate the presence of an adenosine/IL‐6 inflammatory axis in the bronchoalveolar milieu of airway‐compromised horses. While A_2BAR is the predominant proinflammatory AR subtype expressed, A_2AAR appears to modulate inflammatory signalling (IL‐6 expression) by adenosine. Potential relevance: This study supports selective AR targeting as a potential therapeutic approach for the modulation of inflammation in the equine lower respiratory tract.     

细胞因子的应用研究进展

本文综述和分析了动物细胞因子的基因克隆及其在畜牧兽医上的应用和对疾病的治疗，特别是有关IFN、IL、TNF、GM-CSF等几种细胞因子的研究进展及广泛应用，在免疫佐剂、免疫治疗剂、构建基因工程疫苗、基因导入疗法、基因工程药物等的应用上显示了广泛的前景。     

Immunotherapy with autologous tumour antigen-coated microbeads (large multivalent immunogen), IL-2 and GM-CSF in dogs with spontaneous B-cell lymphoma

Cytotoxic T-lymphocyte responses to subcellular antigens are enhanced when antigens are presented on cell-sized silica microbeads called large multivalent immunogens (LMIs). LMIs prepared with tumour cell membrane fragments have induced partial remissions in humans with melanoma and renal cell carcinoma. The purpose of this phase I study was to evaluate the safety of LMIs, prepared with autologous lymphoma cell membranes, along with subcutaneous interleukin 2 (IL-2) and granulocyte-macrophage colony stimulating factor (GM-CSF) in dogs with untreated B-cell lymphoma. After lymph node excision and induction chemotherapy, five dogs were vaccinated with three weekly doses of LMI alone; five with LMI and subcutaneous IL-2 and five with LMI, IL-2 and GM-CSF. No significant toxicity was noted, treatment did not adversely affect disease-free interval and half of the dogs showed measurable delayed-type hypersensitivity reactions to intradermal challenge with LMI, suggesting specific cell-mediated immunity.     

LFA—3和IL—2对机体免疫增强作用

以绵羊红细胞膜提取的ＬＦＡ－３和猪外周血单个核细胞所产生的ＩＬ－２，同时或分别与新城疫Ⅳ系疫苗胸肌注射２２日龄雏鸡，测定鸡体ＨＩ抗体效阶及Ｅｔｇ花环形成率的变化，结果表明，ＬＦＡ－３组抗体效价与对照组差异极显著，ＩＬ－２也可显著提高ＩＶ系苗免疫后ＨＩ抗体效价，且两者的作用具有剂量依赖性。     

Trichostrongylus colubriformis induces IgE-independent CD13, CD164 and CD203c mediated activation of basophils in an in vitro intestinal epithelial cell co-culture model

Gastrointestinal nematodes pose a major risk to the farming of small ruminants worldwide. Infections are typically controlled by anthelmintics, however as resistance to anthelmintics increases, it is necessary that the mechanism of host responses are understood in order to develop alternative control options. It is hypothesised that basophils are involved in the initiation of an anti-parasite immune response, independent of IgE. In this study, the in vitro activation states of CD203c⁺ basophil-like KU812 cells were determined in the presence of Trichostrongylus colubriformis parasitised HT29 epithelial cells with or without mucin. Cell surface expression of CD164, CD107a and CD13 antigens on gated CD203⁺ cells were determined and qRT-PCR was used to examine gene expression changes of IL33 (a Th2 cytokine) and the high affinity IgE receptor (FcɛRIα) within the co-culture. When KU812 basophils encountered T. colubriformis and/or mucin in a parasitised epithelium, the basophils increased cell surface expression of CD13 and CD164 antigens, independent of IgE. T. colubriformis also increased the number of CD203c⁺ KU812 cells that expressed CD13 and CD164 antigens. These data support the in vivo observations of T. colubriformis primary infections in guinea pigs and sheep.     

鸡的7种细胞因子的一些分子生物学特性

细胞因子是免疫系统的重要调节因子 ,能够影响免疫应答的类型和水平。近年来 ,禽细胞因子研究取得很大进展 ,随着更多鸡细胞因子基因的发现及其生物学特性的研究 ,临床应用细胞因子成为可能。文章对 7种鸡细胞因子——干扰素 (I型和 II型 ) ,白细胞介素 (IL-2、IL-6、IL-1 5和 IL-1 8)和鸡髓细胞生长因子(c MGF)一些生物学特性作了综述 ,主要包括c DNA克隆的方法、c DNA及其编码蛋白的生物学特性、检测方法、主要生物学功能 ,并与相应的哺乳动物细胞因子比较 ,阐明鸡的细胞因子与哺乳动物的异同点。     

白细胞介素-1α对仔猪睾丸间质细胞睾酮分泌的影响

用体外培养的仔猪睾丸间质细胞，研究了白细胞介素－１α（ＩＬ－１α）对间质细胞睾酮分泌的影响。结果，ＩＬ－１α能抑制ｈＣＧ诱导的间质细胞分泌睾酮，其睾酮产生量随ＩＬ－１α剂量的增加及作用时间的延长而下降；用２０Ｕ／ｍＬ和５０Ｕ／ｍＬ的ＩＬ－１α作用４８ｈ，可分别抑制６０％及６５％的睾酮分泌量；但无ｈＣＧ诱导时，５０Ｕ／ｍＬ的ＩＬ－１α可使间质细胞基础睾酮的生成量增加１倍。提示睾丸内ＩＬ－１α可能通过多种途径调节间质细胞睾酮的产生。     

间歇光照对肉鸡腹水征及生产性能的影响

４００只７日龄艾维茵肉鸡,随机均分为１,２,３和４组,４个小组垫料平养。１组和３组采用２３Ｌ∶１Ｄ的连续光照程序（ＣＬ）,２组和４组为１Ｌ∶３Ｄ的间歇光照程序（ＩＬ）。１组和２组空气中氨气浓度≤１０ｐｐｍ,３组和４组空气中氨气浓度控制在２５～３５ｐｐｍ。结果表明,间歇光照可显著降低由氨气浓度过高引起的肉仔鸡腹水征和心肺功能异常造成的死亡率,与连续光照相比,间歇光照不影响肉鸡出栏时的体重,但能改善饲料报酬。要想充分发挥间歇光照对肉鸡的有利作用,必须为其提供良好的饲养条件。     

白细胞介素18生物学作用及其在病毒感染中的表达调控

白细胞介素18（IL－18）作为一种能诱导产生IFN－γ的因子，在机体产生抗微生物免疫中起着重要作用。巨噬细胞是生物活性IL－18的主要来源，它们持续转录IL－18的mRNA并表达其前体蛋白。微生物感染可提高巨噬细胞中IL－18基因的表达水平，IL－18分泌的调节以IL－18前体蛋白翻译后的调节为主，而不是对转录活性进行调控。本文综述了IL－18的生物学特性，总结了IL－18促进炎症和免疫调节功能的最新知识，重点讨论病毒感染时巨噬细胞产生IL－18的Caspase依赖性调控。     

腐蹄病节瘤拟杆菌纤毛蛋白与绵羊IL-2融合基因的分泌型表达

用改进的 Mg Cl2 法制备宿主菌株PAK/ 2 pfs感受态细胞 ;由 JM10 5工程菌中提取纤毛蛋白与绵羊白细胞介素 2 ( Pilin-IL-2 )融合基因表达载体 ,转化宿主细胞 PAK/ 2 pfs,筛选出具有 Pilin-IL-2融合基因表达载体的阳性克隆菌株 ;在营养肉汤中进行 Pilin-IL-2融合基因的表达。培养 16～ 18h后 ,离心 ,向上清液中加入饱和 ( NH4) 2 SO4提取 Pilin-IL-2融合基因表达蛋白。用羊抗兔 E型节瘤拟杆菌抗血清与提取的融合蛋白进行对流免疫电泳 ,证明该融合蛋白具有特异性     

Plasma interleukin-6 response is predictive for severity and mortality in canine systemic inflammatory response syndrome and sepsis

BACKGROUND: Sepsis is still a major cause of death in both human and veterinary medicine. Early diagnosis is essential for appropriate treatment. Identification of patients at risk for developing sepsis is already possible in human medicine through the measurement of plasma interleukin-6 (IL-6) levels. In veterinary medicine, however, this has been investigated only in canine experimental models. OBJECTIVES: The purpose of this study was to measure IL-6 plasma levels in dogs with naturally occurring systemic inflammatory response syndrome (SIRS) and sepsis and to analyze the value of IL-6 as a predictive parameter for severity and mortality. METHODS: Included in the study were 79 dogs that had been admitted to the small animal clinics of Munich and Berlin from July 2004 to July 2005 and that satisfied the diagnostic criteria for SIRS and sepsis as defined using established parameters. Measurement of plasma IL-6 levels on days 0, 1, and 2 was performed by the use of a colorimetric bioassay based on IL-6-dependent cell growth. RESULTS: Septic foci were identified in 43 patients (septic group), and 36 patients were enrolled in the SIRS group. The frequency of positive blood cultures was 11%. The overall mortality rate was 48%. Higher plasma IL-6 levels on the day of admission were significantly correlated with a more severe degree of disease, increased mortality rate, and earlier fatality. CONCLUSIONS: Plasma IL-6 concentration is predictive of outcome in canine SIRS and sepsis and may be a valuable laboratory parameter for assessing critically ill dogs.     

运用多座位外显方差法分析中国荷斯坦牛CXCR1和IL8基因与乳房炎易感性的关系

旨在运用多座位外显方差分析法(MPVA)分析中国荷斯坦牛CXCR1和IL8两个基因多个突变位点多态性与乳房炎易感性的交互作用.本研究以南方某大型奶牛场634头(其中102头患乳房炎)中国荷斯坦牛为试验材料,采用PCR-SSCP和直接测序法分析CXCR1-5′端和编码区及IL8 2789/2862内含子3、外显子4和5,2个基因共7个SNPs位点的遗传多态性,并用MPVA法分析CXCR1和IL8基因各位点突变及其基因型组合对奶牛乳房炎易感性的影响.结果,MPVA分析发现CXCR1-1830、CXCR1-1768和IL8 2789/2862三位点交互作用对奶牛乳房炎易感性的影响达到显著水平.CXCR1-1830、CXCR1-1768和IL8 2789/2862三位点交互模型作为奶牛乳房炎易感性相关的基因联合选择模型最佳,MPVA可用于奶牛乳房炎易感性多基因分析模型.     

中国荷斯坦牛IL8基因SNP-233（G＞A）不同基因型的表达差异研究

[目的]本研究旨在研究中国荷斯坦牛IL8基因-233（G＞A）位点不同基因的表达差异水平,为探讨该位点在抗奶牛乳房炎作用提供理论依据.[方法]本试验运用SYBRGreen实时荧光定量PCR技术进行定量测定.[结果]GG基因型个体mRNA的表达量显示高于GA和AA基因型个体的表达量.[结论]IL8基因-233（G＞A）位点对中国荷斯坦牛乳房炎抗性有一定影响,可用于中国荷斯坦牛的抗乳房炎的分子标记辅助选择.     

鸡IL-2在新城疫病毒F基因疫苗免疫中的作用

利用国内首次克隆成功的鸡 IL- 2基因真核表达质粒与新城疫病毒 (NDV) D2 6株 F基因真核质粒 (F基因疫苗 )联合免疫 SPF雏鸡 ,观察了其诱导的免疫应答及免疫保护作用 ,以及不同免疫方式对其免疫作用的影响。结果证实了国内克隆鸡 IL- 2基因的免疫增强作用 ,以及作为免疫佐剂应用于禽类基因免疫的可行性 ,为鸡 IL- 2这一新型佐剂的实际应用提供了重要的试验依据。     

Autologous conditioned serum: the comparative cytokine profiles of two commercial methods (IRAP and IRAP II) using equine blood

Reasons for performing study: Osteoarthritis (OA) is one of the most prevalent and debilitating conditions affecting the horse. Autologous conditioned serum (ACS), commercially available as IRAP and IRAP II, is a recently developed treatment for OA in which plasma is prepared from venous blood by incubation with glass beads for 24 h. This product has been shown to increase anti‐inflammatory cytokines and growth factors in human blood. However, data for equine ACS preparations are lacking. Objectives: To characterise the protein profiles produced by commercially available ACS systems in equine blood. Methods: Blood was drawn from 5 horses into 6 groups: red top vacutainer (control), IRAP and IRAP II, with and without heparin. Samples were collected 1 or 24 h post draw and analysed for IL‐1ra, IL‐10, IGF‐1, TGF‐β, TNF‐α and IL‐1β using ELISAs. Results: Twenty‐four hour IRAP and IRAP II samples contained significantly higher levels of all cytokines relative to 1 h serum controls. At 24 h, IRAP II contained significantly higher levels of IL‐1ra and IRAP contained significantly higher levels of TNF‐α, compared to 24 h controls. In addition, TGF‐β, IL‐10 and IL‐1β in IRAP and IRAP II sera were similar to 24 h serum controls. The addition of heparin significantly reduced levels of IGF‐1, TNF‐α and TGF‐β, and significantly elevated levels of IL‐1ra. Conclusions: The cytokine profile that IRAP II produced is modestly better than IRAP. Incubation of whole blood in glass tubes stimulated cytokine synthesis, although not as efficiently as IRAP II. Potential relevance: Although high levels of IL‐1ra were found in ACS, elevation of other factors suggests these cytokines play a previously understated role in clinical improvements. Because ACS has been shown to alleviate clinical symptoms of OA, the present study suggests that factors other than IL‐1ra alone might be involved in its clinical efficacy. Species‐dependent elevations of cytokines warrant further investigation and optimisation of the systems appears to be necessary based on the differences between human and equine blood.     

纤毛蛋白与绵羊白细胞介素2融合基因工程疫苗对实验动物的体液免疫应答

将转化节瘤拟杆菌（D.nodosus)纤毛蛋白（Pili）和绵羊白细胞介素2（oviIL2)融合基因表达质粒的工程菌BL－21，在含酸苄青霉素营养肉汤培养基中表达，离心获得菌体沉淀物，裂解后配制成Pili-oviIL2融合基因工程疫苗。用加佐剂和不加佐剂的Pili-oviIL2融合基因工程疫苗分别接种2只健康家兔，21天后接种第2次；定期采血，用对流免疫电泳检测试验兔的体液免疫反应。结果发现，加佐剂和不加佐剂的Pili-oviIL2融合基因工程疫苗免疫兔7天即可产生相应抗体，抗体在免疫血清中可维持6个月以上。进一步试验将不加佐剂的Pili-oviIL2融合基因工程疫苗接种3只健康绵羊，同样21天后接种第2次，定期采血，用对流免疫电泳检测试验绵羊的体液免疫反应。同时用Pili基因工程疫苗接种2只绵羊作对照。结果3只绵羊接种Pili-oviIL2融合基因工程疫苗后，分别于7天和14天产生相应的抗体，而接种Pili基因工程疫苗的绵羊于28天产生相应的抗体；被免疫绵羊血清中的抗体可维持6个月以上。用Pili-oviIL2融合基因工程疫苗接种兔和绵羊的免疫试验表明，Pili-oviIL2融合基因工程疫苗具有较好的体液免疫应答反应，重组OviIL2在融合基因工程疫苗中具有良好的佐剂作用。