基于农牧业产品和生活用水的京津冀地区水足迹时空特征研究

水足迹将实体水消费和虚拟水消费联系起来,能真实地反映水资源利用的空间差异状况。运用自下而上的方法,分别计算了京津冀地区市级、区县尺度2000—2014年水足迹与人均水足迹,并分析其时空变化特征。结果表明:(1)市级尺度上,北京市水足迹、人均水足迹最高,天津市次之。石家庄市水足迹、人均水足迹相对较高,衡水市水足迹、人均水足迹均相对较低。京津冀市级平均水足迹由2000年的35.88亿m~3增长到2014年的50.82亿m~3,天津市、北京市增长幅度最大。(2)区县尺度上,北京市主城六区水足迹、人均水足迹最高,北京市其他市辖区和天津市主城六区、滨海新区水足迹、人均水足迹次之,石家庄市主城区水足迹、人均水足迹也相对较高。北部燕山山区、西部太行山区及衡水市所辖区县水足迹、人均水足迹最低。京津冀区县平均水足迹由2000年的2.30亿m~3增长为2014年的3.16亿m~3。北京市市辖区水足迹增长幅度最大,天津市市辖区次之。(3)水足迹的构成比例从大到小依次是消费虚拟水量、生活用水量和生态环境用水量,消费虚拟水量约占水足迹的90%。京津冀生态环境用水所占水足迹比例总体呈现增加的趋势,北京市生态环境用水所占水足迹的比例高于天津市和河北省。本文对京津冀地区水足迹的计算与时空变化特征分析,以期为研究京津冀地区城镇化对水资源量的胁迫效应提供参考与依据。     

Molecular characterization of avian-like H1N1 swine influenza a viruses isolated in Eastern China, 2011

Currently, three predominant subtypes of influenza virus are prevalent in pig populations worldwide: H1N1, H3N2, and H1N2. European avian-like H1N1 viruses, which were initially detected in European pig populations in 1979, have been circulating in pigs in eastern China since 2007. In this study, six influenza A viruses were isolated from 60 swine lung samples collected from January to April 2011 in eastern China. Based on whole genome sequencing, molecular characteristics of two isolates were determined. Phylogenetic analysis showed the eight genes of the two isolates were closely related to those of the avian-like H1N1 viruses circulating in pig populations, especially similar to those found in China. Four potential glycosylation sites were observed at positions 13, 26, 198, 277 in the HA1 proteins of the two isolates. Due to the presence of a stop codon at codon 12, the isolates contained truncated PB1-F2 proteins. In this study, the isolates contained 591Q, 627E and 701N in the polymerase subunit PB2, which had been shown to be determinants of virulence and host adaptation. The isolates also had a D rather than E at position 92 of the NS1, a marker of mammalian adaptation. Both isolates contained the GPKV motif at the PDZ ligand domain of the 3′ end of the NS1, a characteristic marker of the European avian-like swine viruses since about 1999, which is distinct from those of avian, human and classical swine viruses. The M2 proteins of the isolates have the mutation (S31N), a characteristic marker of the European avian-like swine viruses since about 1987, which may confer resistance to amantadine and rimantadine antivirals. Our findings further emphasize the importance of surveillance on the genetic diversity of influenza A viruses in pigs, and raise more concerns about the occurrence of cross-species transmission events.     

温度、pH及效应剂对高粱PEP羧化酶活性的协同作用

纯化的高粱PEP羧化酶活性随pH升高(pH6.6～8.0)而增大。在G6P和Mal存在下,酶活性仍有随pH升高而增大的趋势,但G6P对酶的激活百分率和Mal的抑制百分率随pH升高而降低。高粱PEP羧化酶活性的最适温度高于40℃、酶的催化效率(V_(max)/K_m)随温度升高而增大。高温下,反应激活能降低,Mal对酶活性抑制百分率亦随温度升高而下降,I_(0.5)值增大,Mal增大K_m(PEP)的效应变小。     

Characterization of extracellular proteins produced by Aeromonas hydrophila AH-1

Aeromonas hydrophila is a ubiquitous Gram-negative bacterium which can cause motile aeromonad septicemia in both fish and humans. A. hydrophila secretes many extracellular proteins associated with pathogenicity and environmental adaptability. In this study, an extracellular proteome map of A. hydrophila AH-1 was constructed. The major extracellular virulence factors were characterized by comparing the proteomes of various deletion mutants with that of the wild type. The results suggested that serine protease was involved in the processing of a toxin and secreted enzymes such as hemolysin, glycerophospholipid-cholesterol acyltransferase and metalloprotease. We also showed that expressions of polar and lateral flagellins were under the control of temperature, FlhA, LafK, and RpoN. In addition, three novel proteins (potential effector proteins including one ExoT-like protein) were revealed to be secreted via the type III secretion system (TTSS) of A. hydrophila AH-1. Another novel finding was the demonstration of a crosstalk between the lateral flagellar system and the TTSS in A. hydrophila. These results showed that proteomics is a powerful tool for characterizing virulence factors. The construction of proteome maps will provide a valuable means of finding potential candidates for developing suitable diagnostics and therapeutics for this emerging pathogen.     

Endostatin gene therapy enhances the efficacy of paclitaxel to suppress breast cancers and metastases in mice

Chemotherapy combined with antiangiogenic therapy is more effective than chemotherapy alone. The aim of this study was to investigate whether endostatin, a potent anti-angiogenic agent, could enhance the efficacy of paclitaxel to combat breast cancer. An expression plasmid encoding mouse endostatin (End-pcDNA3.1) was constructed, which produced intense expression of endostatin and inhibited angiogenesis in the chorioallantoic membrane assay. 4T1 breast tumors were established in BALB/c mice by subcutaneous injection of 1 × 10⁵ 4T1 cells. The End-pcDNA3.1 plasmid diluted in the transfection reagent FuGENE^TM was injected into the tumors (around 100 mm²), and paclitaxel was injected i.p. into the mice. Endostatin gene therapy synergized with paclitaxel in suppressing the growth of 4T1 tumors and their metastasis to the lung and liver. Both endostatin and paclitaxel inhibited tumor angiogenesis and induced cell apoptosis. Despite the finding that endostatin was superior to paclitaxel at inhibiting tumor angiogenesis, paclitaxel was nevertheless more effective at inducing tumor apoptosis. The combination of paclitaxel and endostatin was more effective in suppressing tumor growth, metastases, angiogenesis, and inducing apoptosis than the respective monotherapies. The combinational therapy with endostatin and paclitaxel warrants future investigation as a therapeutic strategy to combat breast cancer.     

基于同源重组的裂殖壶菌遗传转化体系的构建及验证

【目的】裂殖壶菌是一种能高效生产DHA的海洋真菌;基因工程技术已经成功应用在微生物改造和代谢机理研究中,利用基因工程技术对裂殖壶菌进行改造首先需要构建适合裂殖壶菌的遗传转化体系;【方法】本文利用电转化的方法将含有18S r DNA同源重组片段的ble基因导入裂殖壶菌中,通过zeocin抗性平板筛选出阳性菌株,并设计ble基因引物,以裂殖壶菌基因组为模板,进行PCR验证ble基因是否成功结合到裂殖壶菌染色体上。【结果】筛选获得的抗性菌株基因组上确实PCR出ble基因片段,对改造菌株与原始菌株进行发酵培养,发现改造后菌株在生物量、油脂含量、DHA含量及脂肪酸分布等方面和原始菌株基本一致。【结论】抗性基因的插入不会影响菌株的正常代谢,该体系的构建为后续其他外源基因导入奠定基础。     

以葡萄糖为底物一步法合成γ-氨基丁酸整合型重组钝齿棒杆菌的构建

[目的]为了构建一株直接利用廉价的葡萄糖合成γ-氨基丁酸的重组钝齿棒杆菌,将来自于植物乳杆菌γ-氨基丁酸合成途径的关键酶谷氨酸脱羧酶基因(lpgad)在产谷氨酸菌株钝齿棒杆菌中进行整合表达,实现葡萄糖到GABA的一步法生产.[方法]运用PCR技术扩增得到带有tac启动子的谷氨酸脱羧酶基因tacgad.通过重叠PCR的方法获得钝齿棒杆菌精氨酸合成途径关键酶N-乙酰谷氨酸激酶(NAGK)基因内部缺失型基因△argB.利用自杀载体pK18mobsacB构建同源整合载体pK18-△argB::tacgad,以△argB的上下游序列为同源臂,通过两次同源重组将tacgad基因整合到钝齿棒杆菌基因组,同时将NAGK基因argB灭活,利用蔗糖致死基因sacB反向筛选标记筛选得到谷氨酸脱羧酶的重组钝齿棒杆菌C.crenatum △argB::tacgad.重组钝齿棒杆菌以葡萄糖为底物进行发酵,测定GABA含量.[结果]重组菌C.crenatum △argB::tacgad成功表达谷氨酸脱羧酶,同时阻断了精氨酸合成途径对谷氨酸到GABA代谢途径的竞争,粗酶液基本检测不到NAGK活性,发酵液无精氨酸合成.通过96 h发酵,重组菌可积累约8.28 g/L的GABA.[结论]本研究通过将谷氨酸脱羧酶基因定向整合到钝齿棒杆菌精氨酸合成途径的关键酶基因argB内部,成功表达谷氨酸脱羧酶的同时阻断竞争途径精氨酸的合成.本研究为实现直接利用葡萄糖合成GABA的一步法生产奠定了基础.     

Five New C19‐Diterpenoid Alkaloids from Delphinium tianshanicum W.T.Wang

Five new diterpenoid alkaloids, tianshanitines A‐E ( 1  –  5 ), along with ten known compounds ( 6  –  15 ), were isolated from the EtOH extracts of the whole plant of Delphinium tianshanicum W.T.Wang . Their structures were determined based on extensive spectroscopic analyses, including 1D‐ and 2D‐NMR, HR‐ESI‐MS, and the structure of tianshanitine C ( 3 ) was confirmed by X‐ray diffraction analysis. Tianshanitine A ( 1 ) is the first example of natural diterpenoid alkaloid containing a benzoyl group at C(1) position. Tianshanitine B ( 2 ) is a rare natural diterpenoid alkaloid bearing a OH group at C(16) position. Compounds 1  –  5 , 6 , 8 , 10 , 12 and 14 were evaluated for cytotoxicity against HCT116, MCF‐7 and HepG2 human cancer cell lines.     

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.