单核细胞增多性李斯特菌氨基肽酶Lmo1711体外表达及其酶活分析

对单核细胞增多性李斯特菌(简称单增李斯特菌)lmo1711基因编码的氨基肽酶进行克隆表达与纯化,并研究该重组蛋白的体外酶学特性。首先通过生物信息学分析预测Lmo1711与氨基肽酶家族成员的亲缘关系及关键活性位点的保守性。利用SWISS-MODEL模拟预测该蛋白的空间结构;构建Lmo1711原核表达载体并转化入E.coli Rosetta中,诱导表达重组目的蛋白,并利用镍离子亲和层析方法纯化目的蛋白;以氨基酸-对硝基苯胺偶联物为底物,Lmo1711通过水解底物N端氨基酸残基产生游离对硝基苯胺单体,405 nm处检测吸光值对该产物进行检测从而分析Lmo1711的酶学特性。在此基础上系统研究Lmo1711对不同氨基酸残基底物的催化特异性,及不同金属离子对该酶活性的影响。经原核表达纯化获得49.3 kDa的重组Lmo1711蛋白,与预测分子量一致;生物信息学分析推测Lmo1711属于M29氨基肽酶家族,且存在保守关键氨基酸活性位点(Glu250、Glu316、His345、Tyr352、His378、Asp380);酶活分析显示,Lmo1711具有较强的氨基肽酶活性,针对不同底物的结合和催化能力差异较大,对亮氨酸残基的亲和程度最高;Lmo1711氨基肽酶活性具有金属离子依赖性,Co~(2+)、Cd~(2+)、Zn~(2+)等多种金属离子均能显著增强其活性,其中Co~(2+)的激活效应最显著。本试验首次发现并证实,单增李斯特菌Lmo1711属于M29氨基肽酶家族成员,具有较强的催化活性,且对金属离子具有不同程度的依赖性。     

细胞因子基因转导对人乳腺癌细胞MHC抗原及细胞膜糖蛋白表达的影响

为探讨细胞因子基因(人IL-2、IL-6)转导对于肿瘤细胞膜MHC抗原及细胞膜糖蛋白表达调控的影响,本文利用脂质体介导的方法,将含人IL-6、IL-2基因的逆转录病毒载体分别导入人乳腺癌细胞系MCF-7细胞中,采用间接免疫荧光染色流式细胞仪测定法,对基因转导的瘤细胞细胞膜糖蛋白及MHC抗原表达进行测定。结果表明经两种基因修饰的MCF-7细胞MHCⅠ型抗原表达均获得增强,此外,基因转导细胞可程度不同地表现出细胞膜多种糖蛋白表达的变化。提示肿瘤细胞膜抗原及糖蛋白表达的改变可能是细胞因子基因转导影响肿瘤细胞免疫原性的重要结构基础。     

The thymic stromal cell line MTSC4 induced thymocyte apoptosis in a non-MHC-restricted manner

Mouse thymic stromal cell line 4 (MTSC4) is one of the stromal cell lines established in our laboratory. While losing the characteristics of epithelial cells, they express some surface markers shared with thymic dendritic cells (TDCs). To further study the biological functions of these cells, we compared the capability of MTSC4 with TDCs in the induction of thymocyte apoptosis, using thymic reaggregation culture system. Apoptosis of thymocytes induced by MTSC4 and TDCs was measured by Annexin V and PI staining and analyzed by flow cytometry. We found that MTSC4 selectively augmented the apoptosis of CD4^ 8^ (DP) thymocytes. This effect was Fas/FasL independent and could not be blocked by antibodies to MHC class I and class II molecules. In addition, MTSC4 enhanced the apoptosis of DP thymocytes from different strains of mice, which implies that MTSC4-induced thymocyte apoptosis is not mediated by the TCR recognition of self peptide/MHC molecules. In contrast to MTSC4, thymocyte apoptosis induced by TDCs was MHC-restricted. Thus, MHC-independent fashion of stromal-DP thymocyte interaction may be one of the ways to induce thymocyte apoptosis in thymus. Our study has also shown that the interaction of MTSC4 stromal cells and thymocytes is required for the induction of thymocyte apoptosis.     

Smad2条件基因剔除载体的构建及LoxP位点有效性鉴定

Smads家族是最新发现的TGF－β信号转导途径中一个重要的新基因家族,SMAD2属于受体激活的SMADs。Smad2在某些肿瘤中发生突变,是一种可能的肿瘤抑制基因。Smad2基因完全剔除小鼠在胚胎期E6．5天死亡,为了研究Smad2在成体各组织器官及肿瘤发生中的可能作用,构建了Smad2条件基因剔除载体,将LoxP置于Smad2基因组序列C末端功能域两侧,并在组成型表达Cre重组酶的大肠杆菌中检测了LoxP位点的功能,该载体的构建为进行Smad2组织特异性基因剔除研究了奠定了基础。     

RETRACTED ARTICLE: Therapeutic DNA Vaccination as a Repair Strategy Following Spinal Cord Injury

Myelin-derived proteins, such as tenascin-R (TN-R), myelin associate glycoprotein (MAG), oligodendrocyte-myelin glycoprotein (OMgp), and Nogo-A, inhibit the central nervous system regeneration. In this study, the DNA vaccine encoding for oligodendrocyte and myelin-related antigens was employed to attenuate the axonal growth inhibitory properties of myelin in the setting of spinal cord injury. Using a rat spinal cord dorsal hemisection model, the vaccine directed against the inhibitory epitopes of Nogo-A, MAG, OMgp, and TN-R was administered intramuscularly once a week following spinal cord injury, supplemented with local application of specific anti-sera against the four antigens. Anterograde labeling of dorsal column fibers showed active axonal regeneration through the lesion site at the eighth week following the treatment in experimental group but not in control groups. Light microscopic and ultrastructural analysis revealed that vaccination with these myelin-related antigens did not lead to demyelinating disease. OMgp and TN-R levels were down-regulated at the lesion site together with a parallel increase in growth-associated protein 43 levels in the treatment groups. This study reveals the effective approach of a DNA vaccine strategy by attaining the special antibody to direct neutralization of the myelin inhibitors during spinal cord injury.     

Risks of Avian Influenza Transmission in Areas of Intensive Free-Ranging Duck Production with Wild Waterfowl

For decades, southern China has been considered to be an important source for emerging influenza viruses since key hosts live together in high densities in areas with intensive agriculture. However, the underlying conditions of emergence and spread of avian influenza viruses (AIV) have not been studied in detail, particularly the complex spatiotemporal interplay of viral transmission between wild and domestic ducks, two major actors of AIV epidemiology. In this synthesis, we examine the risks of avian influenza spread in Poyang Lake, an area of intensive free-ranging duck production and large numbers of wild waterfowl. Our synthesis shows that farming of free-grazing domestic ducks is intensive in this area and synchronized with wild duck migration. The presence of juvenile domestic ducks in harvested paddy fields prior to the arrival and departure of migrant ducks in the same fields may amplify the risk of AIV circulation and facilitate the transmission between wild and domestic populations. We provide evidence associating wild ducks migration with the spread of H5N1 in the spring of 2008 from southern China to South Korea, Russia, and Japan, supported by documented wild duck movements and phylogenetic analyses of highly pathogenic avian influenza H5N1 sequences. We suggest that prevention measures based on a modification of agricultural practices may be implemented in these areas to reduce the intensity of AIV transmission between wild and domestic ducks. This would require involving all local stakeholders to discuss feasible and acceptable solutions.     

苋科植物化学成分的研究进展

苋科植物有65个属,约1000余种,主要分布在热带、亚热带和温带地区.该科的很多植物具有良好的食用和药用价值,化学成分丰富,含有三萜类、甾体类、黄酮类、生物碱类、色素类和多肽类物质.为推动我国苋科植物资源的更深层次利用和开发,对苋科植物的化学成分研究概况做一综述.     

ROC曲线分析在评价入侵物种分布模型中的应用

生态位模型（ecological niche models,ENMs）已广泛应用于物种潜在分布区预测,ENMs的应用也为外来入侵物种的风险分析提供了重要的定量化分析工具,但如何评价不同模型之间的预测效果成了当今研究的热点问题。本文介绍了受试者工作特征（ROC）曲线分析在评价不同生态位模型预测效果中的应用原理和分析方法,并以一种植物病原线虫-相似穿孔线虫（Radopholus similis）为例,应用ROC曲线分析法对其5种模型（BIOCLIM,CLIMEX,DOMAIN,GARP,MAXENT）的预测结果进行了比较分析。5种模型的ROC曲线下面积AUC（Area Under Curve）值分别为0.810,0.758,0.921,0.903和0.950,以MAXENT模型的AUC值最大,表明其预测效果最好;方差分析结果表明,除GARP与DOMAIN模型之间AUC值差异不显著外,其余各模型之间差异显著。     

半干旱黄土高原地区春小麦地膜覆盖研究概述

在黄土高原半干旱地区春小麦上进行的地膜覆盖试验表明,地膜覆盖通过改善耕层土壤生态环境条件,即通过改善水,热状况,活化土壤养分,对提高水分和养分利用效率,实现粮食增产具有重要作用,但近年来在生产实践和科学实验中发现,不合理的长期全生育期地膜覆盖,因在作物生长前期和中期覆膜作物较不覆膜作物生长好,覆膜作物在生长期水分蒸腾损失严重,土壤水分的蒸散（蒸发＋蒸腾）损失远比不覆膜土壤严重,在作物生长后期降水少或没有补充灌溉时,会产生严重的水分肋迫现象,显著抑制小穗分化和灌浆,最终导致收获得指数和产量下降,同时,地膜覆盖的增产作用在一定程度上是以耗竭土壤肥力,特别是在有机物质为代 的,因此,不正确的地膜覆盖（如全生育期的地膜覆盖）,不仅有时起不到显著的增产作用,而且易造成土壤养分,特别是土壤中硝态氮的累积和损失,肥料利用效率降低,土壤生态条件恶化,下降,难以持续高产,因此在确定地膜覆盖范式时,必须要考虑底墒,作物生育期降水,地膜覆盖的阶段性和氮肥的施用等。