携带EBV-LMP2基因的重组MVA痘苗病毒初步免疫效果

目的 观察携带EBV lmp2基因的重组痘苗病毒疫苗在小鼠体内诱导的LMP2特异性细胞免疫应答效果.方法 使用表达EBV lmp2基因的重组痘苗病毒MVA-Lac-LMP2,采用低（2＆#215;105 pfu/只）、中（2 ＆#215; 106pfu/只）、高（2 ＆#215; 107pfu/只）三个剂量组,分别于0、2周免疫BALB/c小鼠,末次免疫1周后应用IFN-γ ELISPOT方法检测小鼠脾淋巴细胞中EBV LMP2特异性CTL应答水平.结果 MVA-Lac-LMP2能诱导小鼠产生EBV LMP2特异性的CTL应答,其中,中、高剂量组小鼠CTL水平明显高于低剂量组（P≤0.01）,中、高剂量组小鼠诱导的CTL应答水平差异没有统计学意义.结论 MVA-Lac-LMP2能够在小鼠体内诱导EBV-LMP2特异性的细胞免疫,应答水平的高低与免疫剂量有关.     

HIV DNA疫苗与重组腺病毒伴随病毒联合免疫效果的研究

目的 构建含HIV-1B亚型中国株gagV3基因的DNA疫苗及重组腺病毒伴随病毒(rAAV)疫苗，并研究DNA疫苗和rAAV联合免疫的免疫效果。方法 将HIV-1B亚型中国株gagV3基因克隆入真核表达载体pCI-neo上，构建了含HIV-1 gagV3基因的DNA疫苗pCI-gagV3。采用电击法将pCI-gagV3质粒转染p815细胞，用G418压力筛选，得到转入重组质粒的细胞系p815-gagV3，用免疫酶法检测细胞系中HIV-1基因的表达。用该DNA疫苗进行小鼠免疫实验，检测免疫效果；用该DNA疫苗初次免疫，含同样gagV3基因的重组腺病毒伴随病毒rAAV-gagV3加强免疫，采用免疫酶法检测免疫小鼠血清中HIV-1特异性的抗体水平，用乳酸脱氢酶法检测免疫小鼠的HIV-1特异性CTL水平。结果 pCI-gagV3可以在p815细胞中表达HIV-1的基因，免疫BALB／c小鼠后可以在小鼠体内诱发HIV-1特异性的细胞和体液免疫反应。HIV-1特异性抗体滴度为1：20；效靶比为50：1时，CTL平均杀伤率为41．7％。pCI-gagV3与rAAV-gagV3联合免疫并不能明显提高抗体水平，但可以提高CTL反应，效靶比为50：1时，CTL平均杀伤率为61.3％，高于单独用DNA疫苗或重组AAV疫苗免疫后产生的CTL活性。结论 DNA疫苗与重组腺病毒伴随病毒联合免疫可以提高免疫小鼠产生的HIV-1特异性CTL反应。     

人β防御素-2与前列腺癌特异性膜抗原嵌合蛋白真核表达质粒构建及其诱导的小鼠特异性免疫应答

探索人β防御素2 (h BD- 2 )和前列腺特异性膜抗原(PSMA)共表达重组核酸疫苗针对前列腺癌的免疫治疗。研究以pc DNA3.1为载体,构建重组质粒pc DNA3.1/PSMA和pc DNA3.1/h BD- 2 - PSMA,通过RT- PCR和免疫组化检测其表达。免疫小鼠后,进行血清中抗体检测,CD4 、CD8 T淋巴细胞数目测定及CTL 特异性杀伤作用检测。结果显示构建的质粒转染COS- 7细胞后能表达目的基因,免疫小鼠后能在体内持久表达,可以诱导产生特异性抗体,能有效的刺激T细胞增生,诱导特异性CTL 反应。当以h BD- 2作为免疫佐剂时,CTL 活性更强。本研究成功的构建了含PSMA的表达质粒,免疫小鼠可以诱导出有效的体液和细胞免疫,为前列腺癌的免疫治疗奠定了一定的实验基础     

柯萨奇B3病毒结构和非结构蛋白DNA 疫苗诱导小鼠产生免疫应答的研究

目的 制备4种柯萨奇B3病毒（CVB3）结构和非结构蛋白重组质粒DNA疫苗,并探讨其诱导机体产生体液和细胞免疫应答的效果。方法 用基因重组技术构建4种CVB3结构和非结构蛋白重组质粒,将各重组质粒体外转染真核细胞,用Western blot检测表达产物;于BALB／c小鼠后腿胫骨前肌注射免疫,于0、4、8周共免疫3次,100μg/次。免疫后不同时间检测体液和细胞免疫应答指标。结果 4种重组质粒酶切出相应大小的片段,经测序证实为CVB3序列,Western blot证实能够在体外真核细胞中表达。pcDNA3/vp2、pcDNA3/VP1、pcDNA3/2A和pcDNA3/3D均可诱导小鼠产生相应的特异性抗体、细胞毒性T淋巴细胞（CTL）和淋巴细胞增殖反应、迟发型超敏反应（DTH）,并对致死量的CVB3m、CVB5和CVB2攻击具有保护作用,表现为病毒攻击后第3天血中病毒滴度降低,第10天心肌病理变化比对照组明显减轻,且小鼠生存率显著提高。其中以pcDNA/VP1和pcDNA3／3D组保护作用最明显。结论 CVB3结构蛋白VP1和非结构蛋白3D质粒DNA有可能用作CVB DNA疫苗的候选基因,值得进一步深入研究。     

人免疫缺陷病毒Ⅱ型核心蛋白DNA疫苗的实验免疫研究

目的　检测HIV 2核心蛋白DNA疫苗诱导Balb c小鼠免疫应答的能力。方法　将表达HIV 2核心蛋白DNA疫苗质粒pVAXIgag肌注Balb c小鼠 ,分析CD4 、CD8 T淋巴细胞的数量、脾特异性CTL反应、血清中HIV 2的特异性抗体水平。结果　重组质粒pVAXI gag免疫组与空载体pVAXI及PBS对照组相比较差异显著 ,血清抗体滴度及淋巴细胞杀伤效应为P <0 .0 1,脾T细胞亚群的数量为P <0 .0 5。结论　HIV 2核心蛋白DNA疫苗能诱导Balb c小鼠产生特异性细胞免疫应答和体液免疫应答     

犬腺病毒DNA疫苗的免疫实验研究

目的研究犬腺病毒DNA疫苗pVAX1-CpG-Loop刺激机体产生免疫应答的效果。方法采用2种不同的免疫途径和免疫方案免疫Balb/c小鼠;免疫后每周断尾采血,分离血清测定血清IgG抗体效价;采用MTT和CCK-8方法检测免疫小鼠的T淋巴细胞增殖活性,EIA试剂盒测定IFN-γ的浓度。结果所有接种DNA疫苗的小鼠均产生了针对抗原病毒的特异性IgG抗体。细胞学试验提示构建的犬腺病毒DNA疫苗也可诱导小鼠产生细胞性免疫应答,重组质粒-蛋白联合的免疫方案在刺激机体细胞免疫应答方面优于单一重组质粒。结论以上结果提示犬腺病毒DNA疫苗pVAX1-CpG-Loop既能诱导小鼠产生特异性的体液免疫应答,也诱导小鼠产生了细胞免疫应答,对于机体具有一定的免疫保护效果。     

登革3型病毒prM-E和NS1多基因重组质粒DNA免疫原性增强效果观察

目的　观察登革 3型病毒的prM E和NS1基因重组质粒DNA混合免疫对免疫原性的增强作用 ,为登革DNA疫苗混合免疫提供实验依据。方法　将登革 3型病毒的prM E和NS1基因重组质粒DNA分别混合及单独免疫BALB/c小鼠 ,采用中和试验及MTT法检测免疫小鼠血清中和抗体及脾细胞特异性CTL(cytotoxicT lymphocytes)杀伤率。结果　混合重组质粒DNA免疫组与单一prM E基因重组质粒DNA免疫组均在末次免疫后第 14天检测到中和抗体 ,在第 33天达到高峰 ,为 1∶32。在末次免疫后第 4 1天 ,当效靶比为 4 0∶1时 ,混合重组质粒DNA免疫组的特异性CTL杀伤率为 15 % ,而 2个单质粒DNA组分别为 10 .9%和 12 .4 %。结论　重组质粒DNA混合免疫可同时诱发小鼠产生体液免疫和细胞免疫 ,而且细胞免疫应答具有一定的增强作用 ,但没有出现特异性CTL杀伤率的协同增强效果。     

HCV结构区DNA疫苗诱发小鼠特异性细胞免疫反应的研究

目的通过 HCV结构区 DNA疫苗直接免疫小鼠 ,诱发其体内特异性细胞介导的免疫反应的研究 ,为 HCVDNA疫苗的研制打下基础。方法用重组的 p BK- CMV质粒体外转染小鼠骨髓瘤细胞 SP2 / 0 ,建立了能体外表达 HCV结构区蛋白的 SP2 / 0 D细胞系 ,分别用 SP2 / 0和 SP2 / 0 D细胞攻击用空质粒或 p BK- CMV质粒免疫过的 Balb/ c小鼠 ,通过肿瘤形成、肿瘤重量、肿瘤组织淋巴细胞的浸润以及不同免疫组小鼠血清中 IL - 2和 IFN -γ含量来观察小鼠体内 T淋巴细胞的活性。结果用 SP2 / 0和 SP2 / 0 D攻击空质粒或 p BK- CMV质粒免疫过的小鼠 15 d后 ,SP2 / 0 D攻击的 p BK- CMV质粒免疫鼠肿瘤重量 ( 0 .9± 0 .12 ) g明显低于空白质粒免疫组和 SP2 / 0接种组 ( P<0 .0 0 1) ,且该免疫组小鼠血清中 IL- 2和 IFN- γ的浓度明显高于其它免疫组 ;此外 ,用 SP2 / 0 D攻击的 p BK- CMV免疫鼠 ,肿瘤病理切片中可见明显的 T淋巴细胞浸润。结论重组的 HCV结构区 DNA疫苗 ( p BK- CMV)能诱导小鼠体内特异性 T淋巴细胞反应 ,HCV DNA疫苗可能为临床疾病的治疗和预防提供一种新的途径。     

EB病毒潜伏膜蛋白2重组逆转录病毒的构建及表达

目的　寻找在真核细胞中表达Epstein Barr(EB)病毒潜伏膜蛋白 2 (Latentmembraneprotein 2 ,LMP2 )的有效途径 ,研究LMP2蛋白功能及深入探讨其在诱导细胞免疫应答中的作用。方法　将EB病毒LMP2蛋白的基因重组至逆转录病毒载体LXSN上 ,通过脂质体将重组质粒导入PT6 7细胞 ,G418筛选抗性克隆 ,收集含重组病毒的上清 ,将其感染小鼠成纤维细胞NIH 3T3 ,测定病毒滴度 ,提取转染细胞DNA进行PCR鉴定 ,间接免疫荧光法检测外源基因在感染病毒的NIH 3T3中的表达。结果　培养上清的病毒滴度为 5 .8× 10 5 PFU ml,聚合酶链反应 (PCR)结果证实 ,转染细胞的DNA中含有目的基因的特异性片段。免疫荧光结果表明EBV LMP2基因在小鼠成纤维细胞中获得表达。结论　重组逆转录病毒成功地将EB病毒LMP2基因整合到细胞中并得以表达     

埃博拉病毒包膜蛋白GP基因DNA疫苗诱导小鼠产生高滴度中和抗体

目的探讨埃扎伊尔-埃博拉病毒(Zaire Ebolavirus)包膜蛋白(GP)DNA疫苗诱导小鼠产生中和抗体的特性及研制埃博拉病毒DNA疫苗的可行性。方法构建埃博拉病毒包膜蛋白GP真核表达质粒Peak13CD5LGP,采用100μg剂量的质粒免疫小鼠,以纯化的埃博拉病毒包膜蛋白亚单位融合蛋白(GP1-Fc)作为抗原,通过ELISA检测及Western blot验证测定血清抗体滴度及特异性。结果 100μg质粒免疫小鼠3次后,小鼠血清中GP中和抗体滴度达到1∶2 000。结论编码埃博拉病毒包膜蛋白GP的DNA疫苗(pEAK13CD5LGP)能够诱导小鼠产生高滴度和持久的中和抗体,作为埃博拉病毒疫苗具有潜在的应用价值。     

Renal dysplasia and asplenia in two sibs

A family is reported in which two sibs, one male and the other female, both died within 24 hours of birth with enlarged polycystic kidneys. Postmortem histology in the second child showed gross renal dysplasia. In both children the pancreas was enlarged, nodular and cystic but the liver appeared macroscopically normal. In the second child, histological examination confirmed pancreatic fibrosis with cystic dilation of ducts, but showed portal fibrosis with bile duct proliferation in the liver.
This combination of findings is very reminiscent of those in a girl and her brother reported by Ivemark et al. (1959). The children reported here also showed absence or hypoplasia of the spleen, cardiac anomalies and other features of the Ivemark syndrome (Ivemark 1955), a quite different, usually sporadic, congenital disorder. It is suggested that the children described here have a distinct lethal congenital disorder, probably inherited in an autosomal recessive manner.     

Schizophrenia in a North Swedish geographical isolate, 1900–1977. Epidemiology, genetics and biochemistry

Over 200 schizophrenic patients belonging to three major and interrelated pedigree complexes have been investigated over the past 30 years in a North Swedish geographically isolated population, presently numbering about 6,000. An intensive investigation of a number of biochemical correlates and genetic markers in a few selected families belonging to one of the major pedigrees has indicated new strategies for the current research program.
Schizophrenia, as defined operationally, is significantly associated with decreased activities of two enzymes (1) blood platelet monoamine oxidase, (2) plasma dopamine-β-hydroxylase, and (3) with the genetic marker Gc² (group specific antigen). Both enzymes are subject to genetic variation. A positive score for linkage between schizophrenia and low plasma DBH activity has been calculated, but, so far, available data are insufficient for discrimination between linkage and partial contribution of genetically controlled low plasma DBH to the pathogenesis of the disease. Alternatively, both mechanisms could be involved.
As a model for continued research, schizophrenia is explained as based on a double dominant-recessive genotype (Aabb), representing a vulnerability which in about 50 % of cases develops into clinical schizophrenia. It is suggested that the dominant mutation (A) operates on or affects MAO activity, and that the recessive genotype (bb) is instrumental in low variates of DBH activity and very likely such variates within the normal range of physiological variation. Moreover, it is suggested that the combined effects of MAO- and DBH-reduced efficiency on the metabolism of e.g. dopamine could be an essential pathogenic mechanism for the schizophrenic illness which is segregating in this population.     

The dominant diseases of modernized societies as omega-3 essential fatty acid deficiency syndrome: Substrate beriberi

About 1900, modern food selection and processing caused widespread $\text{epidemics}$ of the B vitamin deficiency diseases of beriberi and pellagra which, for genetic reasons, often expressed as different diseases ranging from bowel and heart disease to dermatoses and psychoses. But the B vitamins merely help convert essential fatty acids (EFA) into the prostaglandin (PG) tissue regulators and it now turns out that, through hydrogenation, milling and selection of w3-poor southern foods, we have also been systematically depleting, by as much as 90%, a newly discovered trace Nordic EFA (w3) of special importance to primates and sole precursor of the PG3(4) series, even as a concurrent fiber deficiency increases body demand for EFA. Since substrate EFA is processed by many B vitamin catalysts, an EFA deficiency will mimic a $\text{panh}$ypovitaminosis B, i.e., a mixture of $\text{substrate}$ beriberi and $\text{substrate}$ pellagra resembling vitamin beriberi and pellagra but exhibiting as even more diverse $\text{endemic}$ disease. This would consitute a second stage of the Modern Malnutrition and explain why some workers now hold the dominant diseases of modermized societies to be new, nutritionally based, pellagraform yet lipid-related and to range, once again, from heart disease to psychosis. It is an assumption that our dominant diseases are unrelated to each other or are merely revealed by our diagnostic acumen and therapeutic success; and that hydrogenating millions of tons of food oils annually, to destroy the rancidity producing w3-EFA, is safe for $\text{primates}$. Extensive beriberiform disease is reported here in 32 typical cases taken from medical practice which responds strikingly to linseed oil supplements (60% w3-EFA) in confirmation of identical results in Capuchins.     

What will studies of Fulani individuals naturally exposed to malaria teach us about protective immunity to malaria?

There are an estimated over 200 million yearly cases of malaria worldwide. Despite concerted international effort to combat the disease, it still causes approximately half a million deaths every year, the majority of which are young children with Plasmodium falciparum infection in sub-Saharan Africa. Successes are largely attributed to malaria prevention strategies, such as insecticide-treated mosquito nets and indoor spraying, as well as improved access to existing treatments. One important hurdle to new approaches for the treatment and prevention of malaria is our limited understanding of the biology of Plasmodium infection and its complex interaction with the immune system of its human host. Therefore, the elimination of malaria in Africa not only relies on existing tools to reduce malaria burden, but also requires fundamental research to develop innovative approaches. Here, we summarize our discoveries from investigations of ethnic groups of West Africa who have different susceptibility to malaria.     

'Very sore nights and days': the child's experience of illness in early modern England, c.1580-1720

Sick children were ubiquitous in early modern England, and yet they have received very little attention from historians. Taking the elusive perspective of the child, this article explores the physical, emotional, and spiritual experience of illness in England between approximately 1580 and 1720. What was it like being ill and suffering pain? How did the young respond emotionally to the anticipation of death? It is argued that children’s experiences were characterised by profound ambivalence: illness could be terrifying and distressing, but also a source of emotional and spiritual fulfilment and joy. This interpretation challenges the common assumption amongst medical historians that the experiences of early modern patients were utterly miserable. It also sheds light on children’s emotional feelings for their parents, a subject often overlooked in the historiography of childhood. The primary sources used in this article include diaries, autobiographies, letters, the biographies of pious children, printed possession cases, doctors’ casebooks, and theological treatises concerning the afterlife.     

Guidelines for the Validation and Use of Immunoassays for Determination of Introduced Proteins in Biotechnology Enhanced Crops and Derived Food Ingredients

Recent advancements in agricultural biotechnology have created a need for analytical techniques to determine introduced proteins in crops enhanced through modern biotechnology techniques. These proteins are expressed in plant tissues and may be present in food ingredients. Immunoassays are ideally suited for protein detection and may be used as both quantitative and threshold methods. Microplate ELISA and lateral flow devices are two of the most commonly used immunoassay formats for agricultural biotechnology applications. This paper provides general background information and a discussion of criteria for the validation and application of immunochemical methods to the analysis of proteins introduced into plants and food ingredients using biotechnology methods. It is the result of a collaborative effort of members of the Analytical Environmental Immunochemical Consortium. This collaborative effort represents the combined expertise of several organizations to reach consensus on establishing guidelines for the validation and use of immunoassays. Further, the paper offers developers and users a consistent approach to adopting the technology as well as aid in producing accurate and meaningful results.     

HLA in North Colombia Chimila Amerindians

HLA-A,-B,-C,-DRB1 and -DQB1 alleles have been studied in Chimila Amerindians from Sabana de San Angel (North Colombian Coast) by using high resolution molecular typing. A frequent extended haplotype was found:HLA-A*24:02-B*51:10-C*15:02-BRB1*04:07-DQB1*03:02 (28.7%) which has also been described in Amerinndian Mayos Mexican population (Mexico, California Gulf, Pacific Ocean). Other haplotypes had already been found in Amerindians from Mexico (Pacific and Atlantic Coast), Peru (highlands and Amazon Basin), Bolivia and North USA. A geographic pattern according to HLA allele or haplotype frequencies is lacking in Amerindians, as already known. Also, five new extended haplotypes were found in Chimila Amerindians. Their HLA-A*24:02 high frequencies characteristic is shared with aboriginal populations of Taiwan; also, HLA-C*01:02 high frequencies are found in New Zealand Maoris, New Caledonians and Kimberly Aborigines from Australia. Finally, this study may show a model of evolutionary factors acting and rising one HLA allele frequency (-A*24:02), but not in others that belong to the same or different HLA loci.     

Ultracryotomy: development and application (with examples from the yeast cytology) (author's transl)

The preparation steps usually necessary for obtaining ultrathin frozen sections of biological material (chemical prefixation, enclosing, cryoprotective treatment, freezing, sectioning, and post-staining the sections for transmission electron microscopy) are submitted to a critical analysis. The application of cryo-ultramicrotomy, in particularly for cytochemical purposes, is reviewed. Fundamental considerations of chemical prefixation and poststaining are supported by examples from yeast cytology. Furthermore, the efficiency of the cryo-ultramicrotomy (electron optical resolution of ultrastructural details) is demonstrated on yeast cells and protoplasts.     

The universal muscular chamber. A basic unit of the genitourinary, cardiovascular, gastro-intestinal, skeletal, cutaneous, respiratory and other systems, endocameral hypertension; and spasm, the key to their idiopathic diseases and arthropathies

Starting with the integument, we see many organs are contractile sacs or multiples thereof, which tubes or bags constitute the major part of the entire body. Recognition of this basic unit and its characteristics sheds new light, individually and collectively, on many disorders previously considered unrelated. Muscular tears and perforations develop in the walls of these chambers, being no way peculiar to those organs, wherein, hydrochloric acid occurs. So, it is not necessary to explain the absence of excessive acid from patients who exhibit holes in the gastric, uterine, aortic, duodenal, rectal, pulmonary, retina, and other walls. Muscle, not acid is the great common factor relating idiopathic disorders in the gastrointestinal tract to each other and to similar diseases in other systems. When the units are linked together, the lesions tend to appear as arthropathies, i.e. at the joints. Rephrasing common-place observations, frees us from conventional, conceptual cul-de-sacs. An observation is only as good as its interpretation, so all possibilities must be considered, otherwise, we will remain blinded by our misconceptions.