汉滩病毒嵌合基因G2S0.7重组腺病毒免疫学特性研究

对汉滩病毒糖蛋白 (GP)和核蛋白 (NP)的嵌合基因G2S0 .7重组腺病毒的免疫学特性进行研究。将汉滩病毒含有G2S0 .7嵌合基因的重组腺病毒直接免疫Balb/c小鼠 ,用ELISA、微量细胞培养中和实验及淋巴细胞增殖实验检测免疫应答效果 ,以研究嵌合基因的免疫效果。结果用该重组腺病毒免疫的小鼠 ,可诱导产生抗汉滩病毒NP及GP特异性的抗体 ,抗体效价分别为 1 :1 6 0及 1 :2 0。同时重组腺病毒还可刺激机体产生低水平的中和抗体和明确的淋巴细胞增殖反应。说明汉滩病毒嵌合基因G2S0 .7重组腺病毒 ,既可刺激机体产生特异性的抗汉滩病毒体液免疫应答 ,也可刺激机体产生特异性的细胞免疫应答 ,本研究为汉滩病毒基因工程疫苗的研究奠定了实验基础     

中华鳖嗜水气单胞菌微球缓释疫苗的研制及免疫效果

为通过免疫防治途径控制嗜水气单胞菌引起的多种中华鳖疾病,采用复乳化-溶剂挥发法制备缓释微球疫苗,通过灌胃方式免疫中华鳖(体质量150～200 g),每只灌胃0.05 g或0.1g,微球含灭活菌约为6.75×1010/g.免疫应答反应结果显示,缓释微球疫苗体外释放可持续近40 d,血清抗体效价、白细胞吞噬率、白细胞吞噬指数及淋巴细胞转化率最高分别可达512,71.25％,1.51和77.25％.结果表明:中华鳖嗜水气单胞菌微球缓释疫苗可以达到注射灭活菌液疫苗的水平,且持续性略优于注射免疫.通过灌胃方式(0.1 g/只)、注射方式分别对中华鳖进行免疫,研究攻毒后的免疫保护率,结果显示注射免疫组和微球疫苗免疫组的免疫保护率均为85％.免疫应答实验和攻毒实验结果分别证明了中华鳖口服缓释微球疫苗免疫的持续性和有效性.     

DNA疫苗及其在卵黄抗体制备中的应用

DNA疫苗技术由于其独特的作用和优势,现已成为最有发展潜力的新兴免疫技术之一。将DNA疫苗技术和卵黄抗体技术联合起来,可获得效价更高、亲和性更好和成本更低的卵黄抗体。就DNA疫苗和卵黄抗体的各自特点以及DNA疫苗技术在卵黄抗体制备中的应用优势进行综述。     

鸭不同MHC单倍型对禽流感重组疫苗免疫效果的影响

目的不同品系的鸭对禽流感(AI)疫苗诱导的效价有差异,分析这种差异是否与主要组织相容性复合体(MHC)有关。方法用表达H5亚型禽流感HA基因重组鸭瘟活载体疫苗免疫抗原提呈相关转运体TAP基因等位基因型不同的4种SPF鸭,比较其血清HI动态变化。结果免疫后连续观察的9周中,B3群SPF鸭HI抗体效价始终显著高于另外3个群。结论鸭的不同MHC单倍型可能与AI疫苗的不同免疫效力相关。     

微小隐孢子虫表面抗原CP23DNA疫苗免疫山羊诱导免疫应答及保护性研究

观察微小隐孢子虫（Cryptosporidium parvum)子孢子表面抗原CP 23 DNA疫苗免疫山羊诱导其产生免疫应答和保护性作用。将重组质粒pCR3.1-23经鼻粘免疫怀孕山羊，用ELISA测定IgG和IgA抗体滴度，用C.parvum卵囊经口对其后代攻虫感染。抗CP23抗体存在于免疫山羊的血浆和初乳中，且抗体滴度随免疫时间延长而增高。C.parvum卵囊经口感染后代，试验组山羊后代与对照组相比，卵囊排出数量减少，排出时间缩短，微小隐孢子虫表面蛋白CP 23 DNA疫苗能诱导山羊产生特异性免疫应答并对其后代有一定免疫保护作用。     

恒河猴犬瘟热病毒基因疫苗制备单克隆抗体的研究

从感染犬瘟热病毒死亡的恒河猴肝脏组织中提取总RNA,经RT-PCR扩增H全基因并克隆到真核表达质粒pVAX中,构建了基因疫苗pVAX-H。用制备的基因疫苗免疫BaLb/c小鼠,小鼠可以产生免疫应答,免疫鼠脾细胞与SP2/0骨髓瘤细胞融合后,经间接ELISA法筛选,获得5株稳定分泌抗CDV H蛋白的单克隆抗体的杂交瘤细胞株。该5株单抗对应不同的病毒抗原表位,且均不与CPV和MV发生反应, McAb诱生的腹水可体外中和犬瘟热病毒,其中2株中和效价大于1:256。结果表明,CDV H基因疫苗可制备具有一定中和效价的抗CDV的单克隆抗体。     

新城疫Ⅱ系、Ⅰ系疫苗早期免疫的探讨及其免疫应答的形态学观察

试验对具有母源抗体的雏鸡采用新城疫Ⅱ系苗滴鼻首免,20日龄Ⅰ系苗饮水二免,通过对1000羽试验雏鸡血清HI抗体效价的连续检测和攻击强毒试验结果,证明雏鸡首免和二免后,HI抗体效价维持在较高水平,对强毒攻击具有坚强的保护力,至100日龄时保护率仍可达80％,而对照鸡在30日龄时保护率即降至0。Ⅰ系苗饮水二免后,经80天观察,未见鸡群出现任何明显不良反应,二免后3天采取免疫鸡和同日龄同群感染鸡的脑,肺脏、脾脏,腔上囊和血液分别作成匀浆接种9日龄鸡胚,未能分离到Ⅰ系病毒。关于新城疫苗免疫应答的形态学变化,国内外尚乏报道,本试验作了比较系统的观察,注意到雏鸡首免和二免后各个时期的免疫器官:胸腺,腔上囊,脾脏,盲肠扁桃体,肠道淋巴组织和气管,肺脏,肝脏及肾腔等器官的免疫应答表现,特别在Ⅰ系苗二免后,更为明显,免疫后淋巴母细胞和浆细胞的增多反应了HI抗体效价提高的相关免疫状态,气管和肺脏次级支气管上皮下淋巴细胞的大量集聚和形成淋巴小结是另一突出特点,提示采用滴鼻和饮水免疫途径可以诱导呼吸道的局部免疫应答和增强呼吸道的局部抵抗力。试验表明:1日龄雏鸡Ⅱ系苗滴鼻首免,20日龄Ⅰ系苗饮水二免,效力可靠,安全,方法简便易行,具有一定的实用价值,并在形态上对免疫效力予以佐证,结合文献论证了呼吸道局部免疫力在抵抗新城疫强毒感染中的重要意义。     

恒河猴犬瘟热病毒基因疫苗单克隆抗体的制备

从感染犬瘟热病毒死亡的恒河猴肝脏组织中提取总RNA,经RT-PCR扩增H全基因并克隆到真核表达质粒pVAX中,构建了基因疫苗pVAX-H.用制备的基因疫苗免疫BaLb/c小鼠,小鼠可以产生免疫应答,免疫鼠脾细胞与SP2/0骨髓瘤细胞融合后,经间接ELISA法筛选,获得5株稳定分泌抗CDV H蛋白的单克隆抗体的杂交瘤细胞株.该5株单抗对应不同的病毒抗原表位,且均不与CPV和MV发生反应,McAb诱生的腹水可体外中和犬瘟热病毒,其中2株中和效价大于1∶256.结果表明,CDV H基因疫苗可制备具有一定中和效价的抗CDV的单克隆抗体.     

汉滩病毒嵌合基因 G150.7 重组腺病毒免疫学特性研究

对本室已构建的汉滩病毒糖蛋白(GP)和核蛋白(NP)的嵌合基因GlS0.7重组腺病毒的免疫学特性进行研究.将汉滩病毒含有G1S0.7嵌合基因的重组腺病毒直接免疫Balb/c小鼠,用ELISA、微量细胞培养中和实验及淋巴细胞增殖实验检测小鼠的免疫应答效果.用该重组腺病毒免疫的小鼠,可诱导产生抗汉滩病毒NP及GP特异性的抗体,抗体效价分别为1:320及1:40.同时重组腺病毒还可刺激机体产生低水平的中和抗体和明确的淋巴细胞增殖反应.说明所构建的汉滩病毒嵌合基因G150.7重组腺病毒,可同时刺激机体产生特异性的抗汉滩病毒体液免疫应答及细胞免疫应答.     

三联疫苗对大黄鱼常见细菌性疾病免疫保护的实验研究

采用0.5%福尔马林4℃灭活和65℃热灭活2种不同的方法制备三联疫苗作为抗原,以多次腹腔注射或浸泡的方式免疫大黄鱼,通过测定受免大黄鱼血清中抗体凝集效价变化并进行攻毒实验以评估保护效果.结果表明:以福尔马林灭活疫苗注射免疫途径效果最佳,其血清抗体效价平均达1536,攻毒后1周内免疫保护率为88.9%;热灭活疫苗注射免疫途径效果也比较明显,其血清抗体效价平均达1024,攻毒后1周内免疫保护率为76.6%;福尔马林和热灭活疫苗浸泡免疫大黄鱼也有一定效果,免疫保护率分别为55.7%和44.6%.     

PA菌毛株菌苗的应用研究

结合国内外近年来生物科学领域中的迅速进展以及个人多年研究多种菌毛株的经验,作者于1984年建成一株“绿脓杆菌MSHA菌毛株”。本菌毛株具有跨菌属的广谱免疫原性。使用此新型菌毛株,1985年制成“绿脓杆菌MSHA菌毛株菌苗”(简称PA菌毛株菌苗)。经基础实验研究、动物实验、志愿者人体验证及临床试用,均获得良好效应。     

治疗性乙型肝炎疫苗研究反思

 “治疗性乙肝疫苗”是以乙肝病毒(HBV)的表面抗原(s抗原)为基础的生物制剂,目的是激发抗s抗原免疫反应,终止HBV慢性感染.HBV的e抗原与s抗原无抗原性关联,对e抗原的反应也与病毒及感染细胞的清除无关,因此以II期临床试验者血清有关病毒e抗原的数据结果,尚不能判断“治疗性乙肝疫苗”是否有效.疫苗的应用是抵御病毒入侵,而治疗性乙肝疫苗是在病毒已经进入人体后应用,在患者肝细胞可能广泛受累的情况下,疫苗一旦打破耐受激发抗s抗原免疫反应,除能清除血清中游离的病毒和s抗原颗粒外,将直接攻击被感染的肝细胞.由于无法估计慢性HBV感染者肝细胞感染程度,所以无法推测免疫反应发生后,免疫病理所致的肝损程度以及相应的风险.在应用上隐含如此风险,是“治疗性乙肝疫苗”走不出实验室的重要因素之一.     

A point mutation at codon 13 of the N-ras oncogene in myelodysplastic syndrome

Patients with a myelodysplastic syndrome (MDS) which has a risk of leukaemic change exhibit a variable clinical course. It has been suggested that the development of leukaemia in patients with MDS may be related to chromosomal abnormalities or genetic alterations: somatic mutation of the N-ras gene is now considered to be a critical step in the genetic basis of human leukaemogenesis. Here we report that DNAs of bone-marrow cells from three out of eight patients with MDS contained an activated N-ras oncogene, as detected by an in vivo selection assay in nude mice with transfected NIH 3T3 cells. Molecular analysis revealed the same single nucleotide substitution at codon 13 in all three transforming N-ras genes. Each of the three patients showed a progression of the disease and a resulting leukaemic change within the following year. Our observation of the mutation at codon 13 in leukaemic cell DNAs from all three cases suggests that activation of the N-ras gene is important in the development of leukaemia in some MDS cases.     

艾滋病疫苗的研究进展

2005年初,在全球六大州19个国家共有35个艾滋病疫苗候选进入了人体早期临床实验。选用的抗原一般均着眼于诱导细胞免疫和体液免疫双重效果。从疫苗的存在形式上看,可供选择的HIV候选疫苗包括下列几种:传统疫苗(灭活疫苗和减毒活疫苗)、合成肽和蛋白亚单位疫苗、DNA疫苗以及活载体疫苗。目前新型疫苗研究策略包括抗原改造加强免疫原性、免疫佐剂协同以提高疫苗免疫反应强度;新型免疫策略包括初免/加强免疫策略和粘膜免疫策略。由于粘膜感染是艾滋病病毒感染的主要途径之一,因而,探索经粘膜免疫是未来疫苗发展的重要方向。中国艾滋病疫苗研究领域的现状是挑战与机遇并存,需要相关领域专家的共同努力,以促进我国艾滋病疫苗研究的发展。     

基因重组α-干扰素治疗慢性粒细胞白血病31例

目的 :观察干扰素治疗慢性粒细胞白血病的疗效。方法 :应用基因重组α干扰素 (IFN α)治疗 31例慢性粒细胞白血病 (CML) ,其中慢性期 (CP) 30例 ,(1例完全缓解病例 ) ,加速期 (AP) 1例。剂量为隔日 30 0万U平均疗程为 9个月 (3～ 4 0个月 )。结果 :单用IFN α治疗CMLCP 5例 ,其中 2例有效 ;联用甲异靛治疗 12例 ,联用羟基脲治疗 14例 ,有效率分别为 91.6 %和 5 7.1% ,与单用甲异靛或羟基脲治疗相比无显著差异。缓解后有 8例单用IFN α维持治持 ,平均持续缓解时间为 6个月。复查Ph+ CML 16例 ,治疗后均未达到完全转阴。结论 :提示IFN α可用于CMLCP和缓解后维持治疗 ,若加大剂量和延长用药时间 ,有可能进一步提高疗效。IFN α对CMLAP无效     

Humoral and cell-mediated immune responses to live recombinant BCG-HIV vaccines

Several viral and bacterial live recombinant vaccine vehicles are being developed to produce a new generation of vaccines against a broad spectrum of infectious diseases. The human tuberculosis vaccine Mycobacterium bovis bacillus Calmette-Guerin (BCG) has features that make it a particularly attractive live recombinant vaccine vehicle. BCG and other mycobacteria are highly effective adjuvants, and the immune response to mycobacteria has been studied extensively. With nearly two billion immunizations, BCG has a long record of safe use in man. It is one of the few vaccines that can be given at birth, it engenders long-lived immune responses with only a single dose, and there is a worldwide distribution network with experience in BCG vaccination. Recently developed molecular genetic tools and methods for mycobacteria have provided the means to introduce foreign genes into BCG. Here we report that a variety of human immunodeficiency virus type 1 polypeptides can be expressed in BCG recombinants under the control of the mycobacterial hsp70 promoter and that the foreign polypeptides produced in BCG can induce antibody and T-cell responses. These results demonstrate that BCG can be used as a live recombinant vaccine vehicle to induce immune responses to pathogen proteins produced by the bacillus.     

鸭疫里氏杆菌疫苗在雏鸭体内诱导细胞免疫的动态研究

本试验选用1日龄仙湖鸭48只,随机均分为6组,其中1～5组为试验组,第6组为对照组.各试验组按不同的免疫次数、首免龄期、接种剂量,用鸭疫里氏杆菌油乳剂灭活疫苗分别作免疫接种,对照组不作接种.各试验组（包括对照组）在免疫前及免疫后每周采血一次,涂片,用酸性α-醋酸荼酯酶（ANAE）反应方法进行染色,用油镜检测计算T、B淋巴细胞百分比值.研究结果表明：无论是2日龄、7日龄或14日龄首免,首次免疫1周后,雏鸭体内都能产生相应的免疫应答,但2日龄免疫应答能力弱,7日龄较强,14日龄最强.两次免疫的细胞免疫效果比一次免疫效果明显.     

Dynamics of chronic myeloid leukaemia

The clinical success of the ABL tyrosine kinase inhibitor imatinib in chronic myeloid leukaemia (CML) serves as a model for molecularly targeted therapy of cancer, but at least two critical questions remain. Can imatinib eradicate leukaemic stem cells? What are the dynamics of relapse due to imatinib resistance, which is caused by mutations in the ABL kinase domain? The precise understanding of how imatinib exerts its therapeutic effect in CML and the ability to measure disease burden by quantitative polymerase chain reaction provide an opportunity to develop a mathematical approach. We find that a four-compartment model, based on the known biology of haematopoietic differentiation, can explain the kinetics of the molecular response to imatinib in a 169-patient data set. Successful therapy leads to a biphasic exponential decline of leukaemic cells. The first slope of 0.05 per day represents the turnover rate of differentiated leukaemic cells, while the second slope of 0.008 per day represents the turnover rate of leukaemic progenitors. The model suggests that imatinib is a potent inhibitor of the production of differentiated leukaemic cells, but does not deplete leukaemic stem cells. We calculate the probability of developing imatinib resistance mutations and estimate the time until detection of resistance. Our model provides the first quantitative insights into the in vivo kinetics of a human cancer.     

Therapy of tuberculosis in mice by DNA vaccination.

Mycobacterium tuberculosis continues to kill about 3 million people every year, more than any other single infectious agent. This is attributed primarily to an inadequate immune response towards infecting bacteria, which suffer growth inhibition rather than death and subsequently multiply catastrophically. Although the bacillus Calmette-Guerin (BCG) vaccine is widely used, it has major limitations as a preventative measure. In addition, effective treatment requires that patients take large doses of antibacterial drug combinations for at least 6 months after diagnosis, which is difficult to achieve in many parts of the world and is further restricted by the emergence of multidrug-resistant strains of M. tuberculosis. In these circumstances, immunotherapy to boost the efficiency of the immune system in infected patients could be a valuable adjunct to antibacterial chemotherapy. Here we show in mice that DNA vaccines, initially designed to prevent infection, can also have a pronounced therapeutic action. In heavily infected mice, DNA vaccinations can switch the immune response from one that is relatively inefficient and gives bacterial stasis to one that kills bacteria. Application of such immunotherapy in conjunction with conventional chemotherapeutic antibacterial drugs might result in faster or more certain cure of the disease in humans.     

LC治疗急性结石性胆囊炎合并胆汁性腹膜炎疗效观察

目的探讨腹腔镜胆囊切除术(LC)与开腹胆囊切除术(OC)对急性结石性胆囊炎合并胆汁性腹膜炎患者机体内炎症性免疫反应的影响.方法分析136例急性结石性胆囊炎患者的临床资料,将入选者随机分成观察组(LC治疗)和对照组(OC治疗)两组,每组68例.比较两组患者的一般资料、临床症状及手术前后的炎性因子水平.结果两组患者的一般资料差异无统计学意义(P0.05).观察组患者的住院时间、术后死亡率及并发症发生率均明显低于对照组(P0.05).术前两组患者的炎性指标水平差异无统计学意义(P0.05);术后两组患者的各炎性指标比较差异具有统计学意义(P0.05).结论与OC相比,LC可有效降低急性结石性胆囊炎合并胆汁性腹膜炎患者的炎性反应,有助于患者快速建立免疫防御系统.