Soluble periplasmic production of human granulocyte colony-stimulating factor (G-CSF) in Pseudomonas fluorescens

Cost-effective production of soluble recombinant protein in a bacterial system remains problematic with respect to expression levels and quality of the expressed target protein. These constraints have particular meaning today as "biosimilar" versions of innovator protein drugs are entering the clinic and the marketplace. A high throughput, parallel processing approach to expression strain engineering was used to evaluate soluble expression of human granulocyte colony-stimulating factor (G-CSF) in Pseudomonas fluorescens. The human g-csf gene was optimized for expression in P. fluorescens and cloned into a set of periplasmic expression vectors. These plasmids were transformed into a variety of P. fluorescens host strains each having a unique phenotype, to evaluate soluble expression in a 96-well growth and protein expression format. To identify a strain producing high levels of intact, soluble Met-G-CSF product, more than 150 protease defective host strains from the Pfēnex Expression Technology? toolbox were screened in parallel using biolayer interferometry (BLI) to quantify active G-CSF binding to its receptor. A subset of these strains was screened by LC-MS analysis to assess the quality of the expressed G-CSF protein. A single strain with an antibiotic resistance marker insertion in the pfaI gene was identified that produced>99% Met-GCSF. A host with a complete deletion of the autotransporter-coding gene pfaI from the genome was constructed, and expression of soluble, active Met-GSCF in this strain was observed to be 350mg/L at the 1 liter fermentation scale.     

Emergy as a Life Cycle Impact Assessment Indicator

Founded in thermodynamics and systems ecology, emergy evaluation is a method to associate a product with its dependencies on all upstream environmental and resource flows using a common unit of energy. Emergy is thus proposed as an indicator of aggregate resource use for life cycle assessment (LCA). An LCA of gold mining, based on an original life cycle inventory of a large gold mine in Peru, is used to demonstrate how emergy can be incorporated as an impact indicator into a process‐based LCA model. The results demonstrate the usefulness of emergy in the LCA context. The adaptation of emergy evaluation, traditionally performed outside of the LCA framework, requires changes to the conventional accounting rules and the incorporation of uncertainty estimations of the emergy conversion factors, or unit emergy values. At the same time, traditional LCA boundaries are extended to incorporate the environmental processes that provide for raw resources, including ores. The total environmental contribution to the product, doré, is dominated by mining and metallurgical processes and not the geological processes forming the gold ore. The measure of environmental contribution to 1 gram (g) of doré is 6.8E + 12 solar‐equivalent Joules (sej) and can be considered accurate within a factor of 2. These results are useful in assessing a process in light of available resources, which is essential to measuring long‐term sustainability. Comparisons are made between emergy and other measures of resource use, and recommendations are made for future incorporation of emergy into LCA that will result in greater consistency with existing life cycle inventory (LCI) databases and other LCA indicators.     

Toward a Carbon Dioxide Neutral Industrial Park

The industrial park of Herdersbrug (Brugge, Flanders, Belgium) comprises 92 small and medium‐sized enterprises, a waste‐to‐energy incinerator, and a power plant (not included in the study) on its site. To study the carbon dioxide (CO₂) neutrality of the park, we made a park‐wide inventory for 2007 of the CO₂ emissions due to energy consumption (electricity and fossil fuel) and waste incineration, as well as an inventory of the existing renewable electricity and heat generation. The definition of CO₂ neutrality in Flanders only considers CO₂ released as a consequence of consumption or generation of electricity, not the CO₂ emitted when fossil fuel is consumed for heat generation. To further decrease or avoid CO₂ emissions, we project and evaluate measures to increase renewable energy generation. The 21 kilotons (kt) of CO₂ emitted due to electricity consumption are more than compensated by the 25 kt of CO₂ avoided by generation of renewable electricity. Herdersbrug Industrial Park is thus CO₂ neutral, according to the definition of the Flemish government. Only a small fraction (6.6%) of the CO₂ emitted as a consequence of fossil fuel consumption (heat generation) and waste incineration is compensated by existing and projected measures for renewable heat generation. Of the total CO₂ emission (149 kt) due to energy consumption (electricity + heat generation) and waste incineration on the Herdersbrug Industrial Park in 2007, 70.5% is compensated by existing and projected renewable energy generated in the park. Forty‐seven percent of the yearly avoided CO₂ corresponds to renewable energy generated from waste incineration and biomass fermentation.     

Life Cycle of Titanium Dioxide Nanoparticle Production

Life cycle impact of emissions, energy requirements, and exergetic losses are calculated for a novel process for producing titanium dioxide nanoparticles from an ilmenite feedstock. The Altairnano hydrochloride process analyzed is tailored for the production of nanoscale particles, unlike established commercial processes. The life cycle energy requirements for the production of these particles is compared with that of traditional building materials on a per unit mass basis. The environmental impact assessment and energy analysis results both emphasize the use of nonrenewable fossil fuels in the upstream life cycle. Exergy analysis shows fuel losses to be secondary to material losses, particularly in the mining of ilmenite ore. These analyses are based on the same inventory data. The main contributions of this work are to provide life cycle inventory of a nanomanufacturing process and reveal potential insights from exergy analysis that are not available from other methods.     

Global macroecology of bird assemblages in urbanized and semi‐natural ecosystems

Aim Despite the increasing pace of urbanization, little is known about how this process affects biodiversity globally. We investigate macroecological patterns of bird assemblages in urbanized areas relative to semi‐natural ecosystems. Location World‐wide. Methods We use a database of quantitative bird surveys to compare key assemblage structure parameters for plots in urbanized and semi‐natural ecosystems controlling for spatial autocorrelation and survey methodology. We use the term ‘urbanized’ instead of ‘urban’ ecosystems as many of the plots were not located in the centre of towns but in remnant habitat patches within conurbations. Results Some macroecological relationships were conserved in urbanized landscapes. Species–area, species–abundance and species–biomass relationships did not differ significantly between urbanized and non‐urbanized environments. However, there were differences in the relationships between productivity and assemblage structure. In forests, species richness increased with productivity; in both forests and open habitats, the evenness of species abundances declined as productivity increased. Among urbanized plots, instead, both species richness and the evenness of species abundances were independent of variation in productivity. Main conclusions Remnant habitats within urbanized areas are subject to many ecological alterations, yet key macroecological patterns differ remarkably little in urbanized versus non‐urbanized plots. Our results support the need for increased conservation activities in urbanized landscapes, particularly given the additional benefits of local experiences of biodiversity for the human population. With increasing urbanization world‐wide, broad‐scale efforts are needed to understand and manage the effects of this driver of change on biodiversity.     

A constitutive expression system for high‐throughput screening

To reduce the amount of consumables and number of pipetting steps in high‐throughput screening, a constitutive expression system was developed that comprises four different promoters of varying strength. The system was validated by the expression of different sucrose phosphorylase enzymes from Leuconostoc mesenteroides, Lactobacillus acidophilus and Bifidobacterium adolescentis in 96‐deep‐ and low‐well plates at three temperatures. Drastically improved soluble expression in mini‐cultures was observed for the enzymes from L. mesenteroides strains by reducing the promoter strength from strong to intermediate and by expressing the proteins at lower temperatures. In contrast, the enzymes from B. adolescentis and L. acidophilus were expressed most efficiently with a strong promoter. The constitutive expression of sucrose phosphorylases in low‐well plates resulted in a level of activity that is equal or even better than what was achieved by inducible expression. Therefore, our plasmid set with varying constitutive promoters will be an indispensable tool to optimize enzyme expression for high‐throughput screening.     

升温诱发高血压大鼠脑梗塞发病的神经内分泌机制

目的：研究气温骤升导致高血压大鼠发生脑梗塞的神经内分泌机制。方法：采用易卒中型肾血管性高血压（RHRSP）模型,放置于人工模拟气温骤升的高温环境中诱发脑梗塞,检测高温刺激前后大鼠ACTH、CORT、TSH、T3、T4的变化。结果：突然升温使生理组大鼠ACTH和CORT水平表现升高的趋势。模型组高血压大鼠CORT、TSH、T3、T4水平在升温中均呈现升高趋势,但是ACTH水平却明显降低（P〈0.05）。升温后发生脑梗塞大鼠的ACTH和T4水平与升温前比明显下降（P〈0.01）,而TSH水平明显高于升温前水平（P〈0.05）,T3水平不变。结论：高血压机体应激反应系统紊乱,甲状腺刺激素和肾上腺皮质激素的异常波动,是突然高温促发高血压机体脑梗塞发病的重要神经内分泌机制。     

Graphene–Cellulose Paper Flexible Supercapacitors

A simple and scalable method to fabricate graphene‐cellulose paper (GCP) membranes is reported; these membranes exhibit great advantages as freestanding and binder‐free electrodes for flexible supercapacitors. The GCP electrode consists of a unique three‐dimensional interwoven structure of graphene nanosheets and cellulose fibers and has excellent mechanical flexibility, good specific capacitance and power performance, and excellent cyclic stability. The electrical conductivity of the GCP membrane shows high stability with a decrease of only 6% after being bent 1000 times. This flexible GCP electrode has a high capacitance per geometric area of 81 mF cm^?2, which is equivalent to a gravimetric capacitance of 120 F g^?1 of graphene, and retains >99% capacitance over 5000 cycles. Several types of flexible GCP‐based polymer supercapacitors with various architectures are assembled to meet the power‐energy requirements of typical flexible or printable electronics. Under highly flexible conditions, the supercapacitors show a high capacitance per geometric area of 46 mF cm^?2 for the complete devices. All the results demonstrate that polymer supercapacitors made using GCP membranes are versatile and may be used for flexible and portable micropower devices.     

胰岛素强化治疗对胃癌围手术期营养代谢的影响：随机对照研究

目的：探讨胃癌围手术期能量及营养物质的代谢特点,研究强化胰岛素治疗对围手术营养代谢的影响。方法：选取胃中、下部癌病理诊断明确并且外科病房ICU住院时间不少于24h的患者64例,取得知情同意后随机分到强化胰岛素治疗（IIT）组血糖控制在4.4~6.1mmol/L,和传统治疗（CIT）组血糖控制在10mmol/L以下;应用CCM营养代谢监测系统测定围手术期静息能量消耗（REE）,呼吸商（RQ）,每公斤体重静息能量消耗（REE/kg）和脂肪氧化比率,应用多频人体生物电阻抗分析仪测定围手术期人体组分的变化及应用稳态模式评估法计算胰岛素抵抗指数（HOMA-IR）。结果：64例病人入选,每组32例,手术创伤引起术后第1、3天REE水平增加约22%和12%,呼吸商降低至0.759和0.791,REE/kg增加28kcal/kg和26kcal/kg,脂肪氧化比率增加至78%和65%,Ln-HOMA-IR明显增加（P〈0.05）;IIT治疗能降低术后第1、3天Ln-HOMA-IR和REE/kg水平;术后人体指标如细胞内液、脂肪组织、蛋白组织、肌肉组织、瘦体组织和体质量较术前水平明显降低（P〈0.05）;IIT能明显减少脂肪组织、蛋白组织和细胞内液的消耗量（P=0.009,t=0.026）。结论：IIT能够有效降低胃癌围手术期胰岛素抵抗程度、降低静息能量消耗的水平和减少脂肪及蛋白质的消耗。