表达鼠精子Cyritestin蛋白的减毒沙门氏菌疫苗株的构建及其鉴定

目的 构建表达鼠精子eyritestin蛋白的可口服减毒沙让氏菌活疫苗株。方法 应用质粒TAZ83／13，将之进行双酶切获得含有成熟cyritestin蛋白cDNA序列的酶切片段。将其插入表达载体yA3149中，构建重组质粒pCFL。将该重组质粒转入鼠沙门氏伤寒杆菌疫苗株X4632和X4550中，以获得重组菌株X4632（pCFL)和X4550（pCFL)。结果 该两种无抗药性的重组菌株在体外营养选择压力下，可较稳定地携带重组质粒传代繁殖。X4550(pCFL)在体内可较稳定地害居于Peyer‘s结，肠系膜淋巴结和脾脏；X4632（pCFL)可较稳定地定居于Peyer‘s结。二者均可表达被抗cyritestin单抗（mAb)识别的重组cyritestin蛋白。口服该疫苗菌株无明显的毒性作用。结论 X4632（pCFL),X4550（pCFL)疫苗株的构建，为研究以cyritestin为靶抗原的精子抗原避孕疫苗打下了基础。     

IL-15真核表达质粒的构建及对乙肝疫苗诱导的体液免疫应答的影响

目的 研究两种不同的IL-15真核表达质粒对乙肝蛋白疫苗诱导的免疫应答的影响。方法：构建IL-15真核表达质粒(简称pIL-15)和含有IL-12信号肽的IL-15真核表达质粒(简称pIL-2s-15)，CTLL-2细胞增殖实验验证两种质粒真核表达产物的生物学活性。将这两种质粒分别与HBsAg共免疫BALB／C小鼠，用ELISA法检测小鼠血清抗-HBs IgG及IgGl、IgG2a亚类的效价。结果：与HBsAg蛋白疫苗共免疫时，pIL-15可使HBsAg诱导的抗-HBsIgG效价升高，显著高于载体pcDNA3．1与HB—sAg共免疫对照组，pIL-2s-15对HBsAg诱导抗-HBsIgC效价没有明显影响。与HBsAg pcDNA3．1组相比，HBsAg pIL-2s-15组和HBsAg pIL-15组诱生的抗HBsIgG2a亚类均升高，但前者IgG2a／IgG1比值最高，与HBsAg pcDNA3．1组相比差别有显著性；HBsAg pIL-15组IgG2a／IgG1比值与HBsAg pcDNA3．1组相比差别无显著性。结论 pIL-15真核表达质粒可增强蛋白疫苗诱导的体液免疫应答，pIL-2s-15真核表达质粒则主要使免疫应答趋向Th1型。     

卡介苗对豚鼠实验性哮喘的预防性治疗作用

目的:观察卡介苗(BCG)对哮喘豚鼠的预防治疗作用。方法:采用31只豚鼠,分为3组进行处理,分别为对照组、卵蛋白(OVA)致敏组和BCG处理组。用OVA(Ⅲ级)致敏豚鼠复制豚鼠哮喘模型。结果:本模型采用10%的OVA致敏,1%的OVA激发,所有动物都表现有不同程度的过敏反应症状。实验动物在接受BCG注射后,表现为以下特点:一是外周血淋巴细胞和单核细胞增加;二是BALF中细胞分类的变化,支气管肺泡灌洗液(BALF)中以淋巴细胞的增加最为明显。 经过OVA致敏的动物BALF和肺组织中嗜酸性粒细胞(EOS)明显增加,BCG不同程度地降低肺组织EOS的气道浸润及减轻OVA致敏豚鼠的气道反应。结论:[HTSS]使用本实验体系BCG可以减轻实验性哮喘的气道炎症反应。     

流感病毒RNA聚合酶蛋白PB1在小鼠中可诱导亚型间交叉免疫保护

目的探索流感病毒RNA聚合酶PB2和PB1亚基作为实验性流感疫苗候选抗原的可能性.方法以复制型质粒（pSCA）为载体分别构建表达甲1型和甲3型流感PB2和PB1的复制型DNA疫苗,免疫小鼠后分别用甲1型流感病毒（A/PR/8/34）进行鼻腔攻击,观察针对不同亚型流感病毒的复制型DNA疫苗的免疫保护效果.结果本实验所构建的复制型DNA疫苗在真核细胞中均可表达外源基因;本实验采用的复制型质粒载体（pSCA）与传统质粒载体（pcDNA3）在诱导小鼠产生抗体方面无差异,并且都诱导了偏向TH1类的免疫反应;表达甲1型和甲3型流感PB1基因的复制型DNA疫苗均可保护小鼠抵御甲1型流感病毒（A/PR/8/34）的攻击.结论表达甲型流感病毒PB1的复制型DNA疫苗能保护小鼠抵御同型和异型流感病毒的攻击,本实验为流感疫苗研究提供新的候选抗原.     

Protective efficacy in chickens, geese and ducks of an H5N1-inactivated vaccine developed by reverse genetics

We generated a high-growth H5N1/PR8 virus by plasmid-based reverse genetics. The virulence associated multiple basic amino acids of the HA gene were removed, and the resulting virus is attenuated for chickens and chicken eggs. A formalin-inactivated oil-emulsion vaccine was prepared from this virus. When SPF chickens were inoculated with 0.3 ml of the vaccine, the hemagglutinin-inhibition (HI) antibody became detectable at 1 week post-vaccination (p.v.) and reached a peak of 10log2 at 6 weeks p.v. then slowly declined to 4log2 at 43 weeks p.v. Challenge studies performed at 2, 3 and 43 weeks p.v. indicated that all of the chickens were completely protected from disease signs and death. Ducks and geese were completely protected from highly pathogenic H5N1 virus challenge 3 weeks p.v. The duration of protective immunity in ducks and geese was investigated by detecting the HI antibody of the field vaccinated birds, and the results indicated that 3 doses of the vaccine inoculation in geese could induce a 34 weeks protection, while 2 doses induced more than 52 weeks protection in ducks. We first reported that an oil-emulsion inactivated vaccine derived from a high-growth H5N1 vaccine induced approximately 10 months of protective immunity in chickens and demonstrated that the oil-emulsion inactivated avian influenza vaccine is immunogenic for geese and ducks. These results provide useful information for the application of vaccines to the control of H5N1 avian influenza in poultry, including chickens and domestic waterfowl.     

Genetic Vaccination of Mice with Plasmids Encoding the NS1 Non-structural Protein from Tick-borne Encephalitis Virus and Dengue 2 Virus

Although there is a safe, inexpensive and efficacious vaccine against yellow fever, vaccination against other flavivirus diseases is less successful. There is no licensed vaccine against dengue fever and current vaccines against tick-borne encephalitis (TBE) and Japanese encephalitis are expensive and require several injections. Furthermore novel vaccines containing only virus envelope proteins may raise fears over antibody mediated enhancement (ADE) of disease. Here we report the successful use of genetic vaccination against TBE in an experimental animal model using a plasmid containing the coding sequence of a non-structural protein (NS1). Such vaccines would provide inexpensive protection against disease, without raising concerns over inducing ADE on subsequent exposure to heterotypic infectious virus. Attempts to generate chaemeric plasmids to protect against both TBE and dengue fever were less successful. Although these chaemeric plasmids directed the synthesis and secretion of the virus NS1 protein normally, no protection was observed against either TBE or dengue fever.     

结核分枝杆菌Ag85B基因疫苗免疫保护作用的初步研究

目的:研究编码结核分枝杆菌分泌蛋白Ag85B的基因疫苗pTB30m和pTB30s对免疫动物的保护作用。方法:以基因疫苗pTB30m和pTB30s肌注免疫BALB/c小鼠。免疫完成6 wk后,用5×105 CFU的MTB H37Rv毒株经小鼠尾静脉攻击感染。同时用尼龙毛柱分离基因免疫BALB/c小鼠的T细胞,并以5×106 T细胞/只小鼠过继免疫正常BALB/c小鼠,立即用105 CFU的MTB毒株经小鼠尾静脉攻击感染。4 wk后分别计数脾脏中的细菌负荷。结果:与生理盐水对照组相比较,pTB30m及pTB30s质粒免疫组BALB/c小鼠脾脏中的细菌负荷均减少,分别为0.645(log10 CFU,P<0.01)和0.839(log10CFU,P<0.001);而空质粒对照组小鼠脾脏中的细菌负荷减少较少。经质粒pTB30m和pTB30s免疫的BALB/c小鼠的T细胞,过继免疫的正常BALB/c小鼠,对攻击感染的MTB H37Rv毒株在脾脏中的增殖具有部分抑制作用。结论:pTB30s免疫的BALB/c小鼠,对MTB H37 Rv毒株攻击的保护作用优于pTB30m质粒免疫,有望进一步用于结核病的防治研究。     

Decreased cell-mediated cytotoxicity against virus-infected cells in systemic lupus erythematosus.

Cell-mediated cytotoxicity, directed against virus-infected tissue culture cells, was studied with peripheral blood mononuclear cells from 11 patients with systemic lupus erythematosus (SLE) and 12 matched, normal subjects in a 51Cr release assay. Baseline (preimmunization) levels of cytotoxicity against target cells infected with influenza A/Victoria, influenza B/Hong Kong, Newcastle disease virus, and herpes simplex virus were significantly decreased in patients with SLE compared to normal subjects (P less than 0.001), although serum antibody levels to the respective viruses were similar in both groups. After intramuscular administration of inactivated influenza A/Victoria vaccine, SLE patients failed to generate elevated levels of cytotoxicity against A/Victoria-infected cells, in contrast to normal subjects. SLE patients responded with levels of serum hemagglutination-inhibition antibody which were similar to those of normal subjects. Thus, SLE patients manifest decreased cell-mediated cytotoxicity against virus-infected target cells, although humoral antibody responses appeared to be intact. Studies of SLE patients with influenza may help to define the role of cell-mediated immunity in the pathogenesis of certain viral infections.     

Single dose inactivated hepatitis A vaccine: Rationale and clinical assessment of the safety and immunogenicity

In comparison with the classical immunisation schedules (0-1-6 or 0-1-12 months) for hepatitis A, a 0- and 12- or a 0- and 6-month schedule would have important advantages by reducing the number of injections and discomfort and increasing scheduling convenience and patient compliance. It would be convenient if a single dose with enough antigen could protect both rapidly and for at least 12 months, when the booster dose would be given. Several clinical trials have been carried out with an inactivated hepatitis A vaccine containing 1,440 ELU. (1 ml), according to a 0–12 and a 0–6 vaccination schedule. This hepatitis A vaccine is safe and well tolerated. It offers a rapid seroresponse: 14 days after a single dose the seroconversion is 88% (95% C.I.: 84.6–90.9). The 0–12 schedule study showed good persistence of hepatitis A virus (HAV) antibodies with a seroconversion rate of almost 95% at month 12. Booster doses given at 6 or 12 months result in a substantial rise in antibody levels; according to these antibody litres, the 1,440 EL. U. vaccine can be expected to confer comparable duration of protection as the 720 EL. U. vaccine, i.e., 10–20 years. Preliminary data show that tinning of the booster may not be critical for the antibody response. In conclusion, the 1,440 EL. U. hepatitis A vaccine is safe, offers rapid seroconversion, and is highly immunogenic. The persistence of HAV antibodies until month 12 allows a certain flexibility in the administration of the booster: month 6 or 12, and a 0–12 or 0–6 schedule can increase the vaccination compliance. copy; 1994 Wiiey-Liss, inc.     

流行性乙型脑炎病毒活疫苗株SA14-14-2基因稳定性研究

目的 通过研究流行性乙型脑炎活疫苗减毒株基因稳定性，从分子水平证实流行性乙型脑炎活疫苗的遗传稳定性。方法分析流行性乙型脑炎活疫苗主种子、工作种子及其相应的疫苗病毒E蛋白基因核苷酸和氨基酸序列，并与其强毒株和基因库中乙脑病毒减毒株(AF15119)比较。结果乙脑活疫苗主种子、工作种子及其相应的疫苗病毒的E蛋白基因核苷酸序列完全相同。这些病毒E蛋白的氨基酸序列与基因库中乙脑病毒弱毒株(AF315119)比较显示第E447位点氨基酸有差异。结论乙脑病毒活疫苗减毒株遗传学特性稳定。