Induction of multiple cytotoxic T lymphocyte responses in mice by a multiepitope DNA vaccine against dengue virus serotype 1

Dengue virus (DENV) is still a major threat to human health in most tropical and subtropical countries and regions. In the present study, a multi‐epitope DNA vaccine that encodes 15 immunogenic and conserved HLA‐A*0201‐, HLA‐A*1101‐, HLA‐A*2402‐restricted CTL epitopes from DENV serotype 1 (DENV‐1) was constructed based on the eukaryotic expressing plasmid pcDNA^TM3.1/myc‐His(?) A. Immunization of HLA‐A*0201, HLA‐A*1101 and HLA‐A*2402 transgenic mice with the recombinant plasmid pcDNA^TM3.1/myc‐His(?) A‐DENV‐1‐Meg resulted in significantly greater IFN‐γ‐secreting T‐cell responses against most (14/15) CTL epitopes than occurred in mice immunized with the empty plasmid pcDNA^TM3.1/myc‐His(?) A. Additionally, the epitope‐specific T cells directed to some epitopes secreted not only IFN‐γ but also IL‐6 and/or TNF‐α. Finally, the induced epitope‐specific T cells also efficiently lysed epitope‐pulsed splenocytes and DENV‐1‐infected splenic monocytes. The present study confirms the immunogenicity of multi‐epitope DENV vaccine, suggesting that it may contribute to the development of a universal DENV vaccine.

     

肺炎克雷伯菌菌毛粘附素克隆表达及其对体外培养细胞的粘附活性

摘要：【目的】肺炎克雷伯菌(K.pn)与宿主细胞的粘附是致病的首要条件,粘附过程主要通过菌毛粘附素MrkD蛋白介导。为了进一步分析MrkD蛋白与宿主细胞间的粘附机制,进一步确定MrkD蛋白的粘附阻断作用。【方法】构建肺炎克雷伯菌菌毛粘附素融合蛋白原核表达质粒pGEX-4T-mrkD,转入大肠杆菌BL21,优化诱导表达条件,表达产物经亲和层析纯化、凝血酶切除融合蛋白GST标签后,进行SDS-PAGE和Western blot鉴定。激光共聚焦显微镜定位MrkD蛋白在宿主细胞上的结合部位;通过粘附活性试验与粘附动力学实验研究了MrkD蛋白的生物活性。【结果】实验得到了分子量为35 kDa的MrkD蛋白,定位了MrkD蛋白在宿主细胞上的结合部位,并证明了MrkD蛋白可以显著影响肺炎克雷伯菌对宿主细胞的粘附力。【结论】本试验首次证实了MrkD蛋白的粘附阻断作用并观察到了其与宿主细胞的作用位点,为研究肺炎克雷伯菌的致病机制,寻找粘附素功能表位奠定了基础。     

采用蛋白质组学技术筛选大肠癌转移相关蛋白

采用对同一亲本来源、不同转移潜能细胞株SW480和SW620的蛋白质表达谱进行双向凝胶电泳和质谱技术分析,并在蛋白质和mRNA水平进行验证,成功鉴定了10个大肠癌转移相关蛋白,其中SW620细胞株表达上调的蛋白质有磷酸甘油酸变位酶1,磷脂酰乙醇胺结合蛋白和高迁移率族蛋白B-1,而热休克蛋白27,膜联蛋白Ⅰ,甲硫腺苷磷酸化酶,切丝蛋白1和表皮型脂肪酸结合蛋白在SW620中表达下调.大多数差异蛋白质功能涉及肿瘤细胞生长、运动、粘附、凋亡等过程,研究结果为阐明大肠癌转移机制及寻找预测大肠癌转移的潜在标志物提供了理论依据.     

Aminoacyl-tRNA formation in the extreme thermophile Thermus thermophilus

Thermophilic organisms must be capable of accurate translation at temperatures in which the individual components of the translation machinery and also specific amino acids are particularly sensitive. Thermus thermophilus is a good model organism for studies of thermophilic translation because many of the components in this process have undergone structural and biochemical characterization. We have focused on the pathways of aminoacyl-tRNA synthesis for glutamine, asparagine, proline, and cysteine. We show that the T. thermophilus prolyl-tRNA synthetase (ProRS) exhibits cysteinyl-tRNA synthetase (CysRS) activity although the organism also encodes a canonical CysRS. The ProRS requires tRNA for cysteine activation, as is known for the characterized archaeal prolyl-cysteinyl-tRNA synthetase (ProCysRS) enzymes. The heterotrimeric T. thermophilus aspartyl-tRNA(Asn) amidotransferase can form Gln-tRNA in addition to Asn-tRNA: however, a 13-amino-acid C-terminal truncation of the holoenzyme A subunit is deficient in both activities when assayed with homologous substrates. A survey of codon usage in completed prokaryotic genomes identified a higher Glu:Gln ratio in proteins of thermophiles compared to mesophiles.     

Karyotyping of comparative genomic hybridization human metaphases by using support vector machines.

BACKGROUND: Comparative genomic hybridization (CGH) is a relatively new molecular cytogenetic method for detecting chromosomal imbalance. Karyotyping of human metaphases is an important step to assign each chromosome to one of 23 or 24 classes (22 autosomes and two sex chromosomes). Automatic karyotyping in CGH analysis is needed. However, conventional karyotyping approaches based on DAPI images require complex image enhancement procedures. METHODS: This paper proposes a simple feature extraction method, one that generates density profiles from original true color CGH images and uses normalized profiles as feature vectors without quantization. A classifier is developed by using support vector machine (SVM). It has good generalization ability and needs only limited training samples. RESULTS: Experiment results show that the feature extraction method of using color information in CGH images can improve greatly the classification success rate. The SVM classifier is able to acquire knowledge about human chromosomes from relatively few samples and has good generalization ability. A success rate of moe than 90% has been achieved and the time for training and testing is very short. CONCLUSIONS: The feature extraction method proposed here and the SVM-based classifier offer a promising computerized intelligent system for automatic karyotyping of CGH human chromosomes.     

腺苷及其类似物对猪血小板肌动蛋白聚合的抑制作用及可能机理

凝血酶和ADP刺激血小板肌动蛋白的聚合,腺苷、5′-氯-5′-脱氧腺苷及2′-脱氧腺苷抑制凝血酶和(或)ADP诱导的肌动蛋白聚合;腺苷和5′-氯-5′-脱氧腺苷对磷脂酰肌醇的磷酸化有抑制作用,且呈剂量效应关系;腺苷对凝血酶刺激的血小板中肌醇二磷酸的生成有抑制作用。本实验提示腺苷及其类似物对肌动蛋白聚合的抑制作用可能与它们对肌醇磷脂转换的抑制有关。     

EB病毒潜伏膜蛋白1在鼻咽癌细胞中通过ERK介导Ets-1表达

为了探讨EB病毒编码的潜伏膜蛋白1(LMP1)对核转录因子Ets-1表达和活化的影响,并证实细胞外信号调节激酶1/2（ERK1/2）参与了该过程,选用可调控表达LMP1的鼻咽癌细胞系L7,应用蛋白质印迹法检测Ets-1、p-ERK蛋白质表达,免疫共沉淀-蛋白质印迹法检测Ets-1磷酸化状态,使用ERK1/2特异性小分子阻断物PD98059作用后,蛋白质印迹法检测p-ERK、Ets-1表达及磷酸化变化.结果显示：在L7细胞中诱导性LMP1可促进p-ERK、Ets-1蛋白质表达及其苏氨酸残基磷酸化,在一定范围呈时间和剂量效应;通过PD98059对诱导性LMP1作用的干预发现,p-ERK大部分表达被阻断,而Ets-1表达及其苏氨酸磷酸化也被部分阻断,以上结果提示ERK部分介导了LMP1诱导Ets-1表达和活化.     

Tissue microarray study for classification of breast tumors

Clinical and pathological heterogeneity of breast cancer hinders selection of appropriate treatment for individual cases. Molecular profiling at gene or protein levels may elucidate the biological variance of tumors and provide a new classification system that correlates better with biological, clinical and prognostic parameters. We studied the immunohistochemical profile of a panel of seven important biomarkers using tumor tissue arrays. The tumor samples were then classified with a monothetic (binary variables) clustering algorithm. Two distinct groups of tumors are characterized by the estrogen receptor (ER) status and tumor grade (p = 0.0026). Four biomarkers, c-erbB2, Cox-2, p53 and VEGF, were significantly overexpressed in tumors with the ER-negative (ER-) phenotype. Eight subsets of tumors were further identified according to the expression status of VEGF, c-erbB2 and p53. The malignant potential of the ER-/VEGF+ subgroup was associated with the strong correlations of Cox-2 and c-erbB2 with VEGF. Our results indicate that this molecular classification system, based on the statistical analysis of immunohistochemical profiling, is a useful approach for tumor grouping. Some of these subgroups have a relative genetic homogeneity that may allow further study of specific genetically-controlled metabolic pathways. This approach may hold great promise in rationalizing the application of different therapeutic strategies for different subgroups of breast tumors.