Cytogenetic stability of chicken T-cell line transformed with Marek's disease virus: atomic force microscope, a new tool for investigation

The Marek's disease virus (MDV) integration may induce a novel organization of chromatin architecture with a modified genetic expression. In our opinion it is worthwhile trying to relate cytogenetic stability to functional modifications. Recently, atomic force microscopy technique was applied to study the structure of chromosomes at a nanoscale level. This high resolution allows to investigate the different structure of chromatin in order to study cytogenetic stability and chromosome aberrations due to MDV insertion. In this paper data are presented indicating a duplication [78,WZ,dup(1p)(p22-p23)] and a deletion [78,WZ cht del(3)(q2.10)] of Chromosomes 1 and 3 relatively. Relationships between GTG (G-bands by Trypsin using Giemsa) bands and the topography of chromosomes are also discussed, naming them Topographic Banding. The architecture of chromosomes observed by AFM can be related to the data obtained with classic banding techniques thus overcoming the optical resolution limits. The presence of chromatin bridges between sister chromatids at most of the heterochromatic regions is also evidenced. Besides, we present different studies of the longitudinal and transversal symmetry of the hetero and euchromatic regions to clearly demonstrate a different underlying architecture of these regions. It is indeed evident that the heterochromatic bands are more symmetrical than euchromatic bands.     

Molecular characterization of major and minor MHC class I and II genes in B21-like haplotypes in chickens

The major histocompatibility complex (MHC) sequences of three B21-like haplotypes deriving from very different origins including the Red Jungle Fowl Gallus Gallus gallus were compared with the MHC sequences of the standard B21 haplotype from Scandinavian White Leghorn Gallus domesticus. The present analysis reveals two cDNA sequences for B-F and two cDNA sequences for B-LB for every B21-like haplotype, including B21 itself. Contrary to expectation, no sequence polymorphism in the antigen-binding domains of the MHC genes, between the investigated haplotypes, was found. The relative level of MHC class I molecules on the surface of leukocytes measured by flow cytometry was also analysed and found to be low in Marek's Disease (MD)-resistant B haplotypes (B21 and B21-like) and high in MD-susceptible B haplotypes (B15 and B19). However, in heterozygous (resistant/susceptible) animals, the relative level was almost as high as in susceptible haplotypes.     

A genome‐wide SNP scan reveals two loci associated with the chicken resistance to Marek's disease

Marek's disease (MD) is a neoplastic disease in chickens, caused by the Marek's disease virus (MDV). To investigate host genetic resistance to MD, we conducted a genome‐wide association study (GWAS) on 67 MDV‐infected chickens based on a case and control design, including 57 susceptible chickens in the case group and 10 resistant chickens as controls. After searching 38 655 valid genomic markers, two SNPs were found to be associated with host resistance to MD. One SNP, rs14527240, reaching chromosome‐wide significance level (P < 0.01) was located in the SPARC‐related modular calcium‐binding 1 (SMOC1) gene on GGA5. The other one, GGaluGA156129, reaching genome‐wide significance (P < 0.05), was located in the protein tyrosine phosphatase, non‐receptor type 3 (PTPN3) gene on GGA2. In addition, expression patterns of these two genes in spleens were detected by qPCR. The expression of SMOC1 was significantly up‐regulated (P < 0.05), whereas the expression of PTNP3 did not show significance when the case group was compared with the control group. Up‐regulation of SMOC1 in susceptible spleens suggests its important roles in MD tumorigenesis. This is the first study to investigate MD‐resistant loci, and it demonstrates the power of GWASs for mapping genes associated with MD resistance.     

人精液对RAJI细胞中EPSTEIN-BARR病毒早期抗原的诱导

在大戟科、瑞香科、豆科和其它科植物中,发现有些植物含有能诱导人伯基特淋巴瘤细胞——Raji细胞中EB病毒早期抗原的物质,并已证明其中有的是很强的促癌物质,如12-0-十四烷酰巴豆醇-13乙酸酯(TPA)、桐油提纯的HHPA、芫花和黄芫花等。某些人的精液也含有能激活EB病毒的物质。本文报告中国人的精液,以及人精液与从宫颈癌病人分离的细菌培养液,对EB病毒早期抗原的诱导作用和协同诱导作用。     

Epstein-Barr病毒基因工程活疫苗人体免疫的初步研究

Epstein-Barr(EB)病毒的原发感染发生在儿童时期,在我国3～5岁儿童的感染率为70％～90％。感染后终生带毒,并经唾液不断排出病毒。我国南方是鼻咽癌高发区,其发病率和死亡率均占恶性肿瘤的第一位。早期诊断方法的改进和早期治疗,使鼻咽癌治疗后的5年生存率明显增加,但不能降低发病率。EB病毒疫苗有可能成为控制该病的有效手段之一。 Epstein等人从淋巴母细胞株(B95-8细胞)细胞表面提取EB病毒膜抗原(MA),用于免疫棉顶猴能产生中和抗体。免疫动物能抵抗EB病毒攻击后所诱发的恶性淋巴瘤。该中和抗体在体外能中和EB病毒的转化活性。EB病毒的主要膜抗原(MA)是由分子量220kD和     

用重组痘苗病毒感染动物细胞表达的Epstein-Barr病毒膜抗原检查IgA/MA抗体

用EB病毒膜抗原基因重组的痘苗病毒感染动物细胞,其细胞表面可表达EB病毒膜抗原,以此膜抗原作为诊断抗原检测血清中IgA/MA抗体,明显优于常用的B95-8细胞表面膜抗原,从而为研究人群血清抗体反应与鼻咽癌的关系,为鼻咽癌的诊断和普查开辟了新的途径。     

EB病毒及其疫苗的研究进展

EB病毒广泛存在于人群中,它的潜伏感染与多种疾病密切相关,包括鼻咽癌等恶性肿瘤,研制疫苗用于EB病毒相关疾病的预防和治疗十分必要。本综述了EB病毒的结构及基因表达特点、EB病毒潜伏期感染基因表达及潜伏期基因产物的功能,以及研制EB病毒疫苗的靶抗原的选择。     

应用纯化的重组Epstein-Barr病毒早期抗原建立检测鼻咽癌病人血清IgA/EA抗体的ELISA方法

利用凝胶和离子交换柱(Mono Q)两次层析,将大肠杆菌表达的EB病毒早期抗原P138片段多肽纯化。以此P138为抗原,增加鼠抗人IgA单克隆抗体以扩大IgA的反应,建立了三步ELISA法。用本法检查了100例鼻咽癌病人和63例正常人血清中抗EB病毒IgA/EA抗体,病人血清的阳性检出率为86％,正常人有3例阳性(4.7％)。此结果表明,三步ELISA法较常用的间接ELISA法(阳性检出率为71％)敏感。     

Foreword: Protein secretion and the Golgi apparatus

Resonance and nonresonance Raman spectra have been obtained from neoplastically transformed and normal avian lymphocytes. The acyl chains of membrane phospholipids of neoplastic cells are more highly unsaturated than those of normal cells. The observation of prominent carotenoid bands in both cell populations indicates the availability of a sensitive, intrinsic probe of membrane potential and local membrane environment.     

应用基因重组Epstein-Barr病毒早期抗原P83蛋白检测鼻咽癌患者血清中的IgA/EA抗体

将pUCB质粒表达的P83蛋白应用于免疫印迹法(IB)和ELISA中,检测了85例鼻咽癌(NPC)患者和100例健康人血清,同时与免疫酶法(IE)作比较。结果表明,免疫印迹法对NPC患者血清阳性检出率为94％;ELISA法阳性检出率为88％;而IE法阳性率为64％。三种方法检测健康人血清出现低水平IgA/EA抗体的阳性率分别为4％、3％及2％。用IE法检测IgA/EA抗体为阴性的NPC患者血清,用IB法检测的阳性率达87％,ELISA法阳性检出率为77％。IB法与ELISA法之间具有较好的正相关(r=0.67,P<0.01)。     

应用蛋白印迹法检测鼻咽癌病人血清中抗Epstein—Barr病毒IgA/EA抗体

曾毅等建立了一系列检测EB病毒IgA/VCA和IgA/EA抗体的鼻咽癌早期诊断方法,取得了满意的结果。为了进一步提高对鼻咽癌诊断更为特异的IgA/EA抗体的检出率,我们建立了检测EB病毒IgA/EA抗体的蛋白印迹法。方法敏感特异,结果令人满意。 本法中所用的两个质粒系由本实验室与西德Pettenkofer研究所Wolf教授的实验室合作构建。pUCARG1140和pUC9MBcE3.2质粒均为表达质粒,前者携带着来源于EB病毒Bam     

表达乙型肝炎病毒表面抗原和Epstein-Barr病毒膜抗原的双价痘苗病毒的组建

从痘苗病毒天坛株分离了晚期11k蛋白编码基因的启动子,以痘苗病毒天坛株为载体,构建了双价的重组痘苗病毒。分别在7.5k和11k蛋白基因启动子的控制下,表达乙型肝炎病毒表面抗原和EB病毒的膜抗原。用重组痘苗病毒免疫的家兔,同时产生对这两种抗原的抗体。免疫电镜下观察到乙型肝炎病毒表面抗原颗粒。     

TLR9 agonists induced cell death in Burkitt's lymphoma cells is variable and influenced by TLR9 polymorphism

Toll-like receptor 9 (TLR9) triggering is a promising novel strategy to combat cancer as it induces innate and adaptive immunity responses. B-cell lymphoma is unique in this context as tumor cells express TLR9 and may harbor latent Epstein-Barr virus (EBV), a gamma-herpesvirus with remarkable oncogenic potential when latent. Latent EBV may be promoted by TLR9 triggering via suppression of lytic EBV. Here, we elaborated an initial assessment of the impact of TLR9 triggering on EBV-positive and EBV-negative B-cell lymphoma using Burkitt''s lymphoma (BL) cell lines as an in vitro model. We show that, independent of the presence of EBV, the TLR9 ligand oligodeoxynucleotide (ODN) CpG-2006 may or may not induce caspase-dependent cell death in BL cells. Moreover, ODN CpG-2006-induced cell death responses of BL cells were associated with TLR9 single-nucleotide polymorphisms (SNPs) rs5743836 or rs352140, which we detected in primary BL tumors and in peripheral blood from healthy individuals at similar frequencies. Thus, our findings suggest that the effect of TLR9 agonists on BL cells should be tested in vitro before installment of therapy and TLR9 SNPs in BL patients should be determined as potential biological markers for the therapeutic response to treatment targeting innate immunity.     

Active EB virus infection in adults: a case series of 8 patients in a single institution and a review of related literature

Adult patients with chronic active EB virus infection (CAEBV) are few; however the disease runs an aggressive course. The goal of this study was to investigate the clinical characteristics, treatment as well as prognosis of CAEBV in adults. The clinical data of eight adult patients with CAEBV of were analyzed retrospectively. There were five male and three female patients with a median age of 47 (25-67) years old. The main clinical manifestations were fever, lymphadenopathy, splenomegaly, liver damage, abnormal coagulation and cytopenia. Peripheral blood EB virus DNA titers were significantly higher in all patients (4.14×10⁴—4.60×10⁵copy/mL). Most patients received treatment with glucocorticoids, cyclosporine and chemotherapy containing etoposide (as recommended in the HLH-2004 program or the scheme of ECOP). One patient progressed to aggressive natural killer (NK) cell leukemia in 5 months after diagnosis. Another patient without fever or cytopenia is still living now, after receiving hormone therapy. The remaining 6 cases died with a median survival of 15 months. In conclusion, CAEBV in adults is a rare but serious disease, often with multiple system damage, from which the prognosis is very poor. Further research is necessary to improve the understanding of this disease.     

Epstein-Barr病毒膜抗原基因免疫的研究

将Ｅｐｓｔｅｉｎ－Ｂａｒ病毒（ＥＢＶ）膜抗原（ＭＡ）ＢＬＬＦ１基因,插入含有ＣＭＶ启动子的真核表达载体ｐｃＤＮＡ３下游ＢａｍＨＩ位点,构建成真核表达质粒ｐｃＤＮＡ３－ＭＡ。将纯化的ＤＮＡ注射Ｂａｌｂ／ｃ小鼠股四头肌。经免疫的动物产生抗ＥＢ病毒ＭＡ特异性的抗体和中和抗体,依赖抗体细胞介导的细胞毒作用（ＡＤＣＣ）,特异性Ｔ淋巴细胞增生性反应及细胞毒性Ｔ淋巴细胞（ＣＴＬ）杀伤作用。基因免疫与基因－蛋白联合免疫效果相似。不同启动子控制下的ＭＡ基因,诱发小鼠免疫应答无明显差异。     

The thyroid "microsomal" antigen is an epitope on the thyrotropin receptor

Antimicrosomal antibodies are present in the sera of most patients with autoimmune thyroiditis, and Graves' disease. It has, in general, been difficult to separate antimicrosomal activity from that directed against the thyrotropin (TSH) receptor in Graves' IgG preparations. The "microsomal" antigen has been localized to the endoplasmic reticulum and microfollicular aspect of thyrocytes; its structure is however unknown. In an attempt to identify the thyroid microsomal antigen, we studied the interaction of Hashimoto's IgG with high microsomal antibody titre and negative for thyroglobulin with purified thyroid plasma and light microsomal membranes. We allowed Hashimoto's, Graves', and control IgGs to bind to protein blots of thyroid plasma membranes resolved on SDS-PAGE under non-reducing conditions. All seven Hashimoto's IgG at a concentration of 2 mg/ml interacted with an M approximately 197,000 polypeptide corresponding to the TSH holoreceptor. By contrast to Graves' IgG (which were positive at 1 mg/ml), however, this binding was not blocked by pretreatment of the protein blots with TSH. Normal IgGs showed no binding at concentrations of up to 2 mg/ml. Both Hashimoto's and Graves' IgG interacted with TSH-affinity column-purified receptor preparations. Two of the Hashimoto's IgGs induced adenylate cyclase activation in thyroid plasma membranes, three inhibited TSH-stimulated enzyme activation, and two were without effect. Two classes of autoantibodies, other than TSH receptor directed, were encountered; one class raised to antigens common to all seven patients and another class unique to individual patients, eg, Mr 210,000 and Mr 20,000 polypeptides. We propose that the TSH receptor has multiple epitopes (functional domains), and the one to which antimicrosomal antibody bind is likely to be spatially separated from that with which Graves' IgG and TSH interact. Differences in affinity or number of sites allows for the demonstration of Graves' IgG against a background of antimicrosomal antibody.     

EB病毒早期蛋白P54的原核表达及其免疫原性的研究

PCR法获得编码EB病毒早期蛋白P54的基因BMRFl,序列分析后亚克隆入原核表达载体pET30a。表达质粒pET30a-BMRF1在大肠杆菌BL21(DE3)菌株中经IPTG诱导后表达了P54抗原,SDS—PAGE表明其相对分子质量为51000;采用镍离子亲和柱纯化重组蛋白。Western印迹结果表明纯化蛋白免疫BALB／c小鼠后产生了P54特异性抗体。间接免疫荧光表明免疫血清可以识别激活的Raji细胞中表达的P54蛋白。以上结果表明构建了原核表达质粒pET30a-BMRF1并在大肠杆菌细胞中成功表达EB病毒早期蛋白P54,表达蛋白具有很好的抗原性和免疫原性。     

The Status of Immunohistochemical Studies in Lymphoma Diagnosis

Immunohistochemical studies of formalin fixed paraffin embedded sections today constitute an essential component of the diagnostic process for malignant lymphoma. Lineage related markers are of value in distinguishing various lymphomas from anaplastic large and small cell carcinomas, and from sarcomas. Other more recently available markers are of value in establishing prognosis and, on occasion, in differentiating between neoplastic and reactive proliferations. Meticulous attention to reagent control and technical procedures is essential to all such studies.     

肝炎病毒与EB病毒重叠感染

为探讨肝炎病毒（ＨＶ）与ＥＢ病毒（ＥＢＶ）重叠感染的状况和后果,我们用免疫酶法对１５４例各型病毒性肝炎患者作了ＥＢＶＩｇＡ抗体检测。结果发现,急性肝炎、慢性轻度肝炎、慢性中度肝炎、肝炎肝硬化、慢性重型肝炎和原发性肝癌ＶＧＡ－ＩｇＡ抗体的阳性率分别为２４．０％、３０．０％、５３．３％、６３．３％、４０．０％和７２．７％,与健康人（５．３％）比较,有非常显著升高（Ｐ＜０．０１）;原发性肝癌又较急性肝炎和慢性轻度肝炎高,并有非常显著意义差异（Ｐ＜０．０１）。ＨＢＶ和ＨＡＶ＋ＨＢＶ感染者比较,前者又较后者低（Ｐ＜０．０１）。重叠感染者的临床表现均为“肝炎型”,未见咽炎、腺热、胃肠、肺炎、肾炎、神经等类型。重叠感染者的ＣＤ＋３及ＣＤ＋４Ｔ细胞下降,ＣＤ＋８Ｔ细胞及ＩｇＧ,ＩｇＭ升高,与健康人比较差异非常显著意义（Ｐ＜０．０１）。结果提示：ＨＶ感染,不仅因免疫失调易感ＥＢＶ,又可因重叠感染而进一步使免疫功能失调;对病毒性肝炎的处理应强调免疫调节治疗。