Expression of bovine beta-lactoglobulin/human erythropoietin fusion protein in the milk of transgenic mice and rabbits

We have generated several transgenic mouse lines and rabbits expressing efficiently (up to 0.3 mg/ml in mice and up to 0.5 mg/ml in rabbits) human erythropoietin in their milk as bovine beta-lactoglobulin fusion protein. Human erythropoietin cDNA was inserted in frame into exon 5 of the bovine beta-lactoglobulin gene with a linker oligonucleotide encoding the cleavage site for bacterial IgA protease. RNA analysis performed on one lactating transgenic mouse and one transgenic rabbit revealed that the fusion gene was expressed almost exlusively in the mammary gland, although low amounts of transgene-derived RNA were detectable in salivary glands and uterus or in the kidney. The fusion protein was specifically cleaved with IgA protease. The erythropoietin part obtained upon digestion had a lower molecular mass than recombinant erythropoietin produced in Chinese hamster ovary cells. By deglycosylation analysis it was shown that the difference in size was due to a different type of glycosylation. Biological activity of the fusion protein, as determined by growth stimulation of TF-1 erythroleukemia cells, was less than 15% of that of human recombinant erythropoietin. Upon digestion of the fusion protein with IgA protease, biological activity comparable to that of the recombinant erythropoietin was recovered. Transgenic males and virgin females did not show signs of enhanced erythropoiesis, but lactating females expressing the transgene displayed transient increases in their hematocrit values.     

Experiments with negative ions in3He superfluids

The interaction between negative ions and ³ He superfluids was studied by using a time-of-flight spectrometer having a 10^–3 relative resolution. The ions guided through a sample of rotating³He-A were found to be focused into the cores of the vortices present in this rotating superfluid. Two different types, with different core structures and presumably different numbers of circulation quanta associated with them, were found. The ion mobility along the vortex cores, _c , observed in the absence of external magnetic field, can be explained by the mobility anisotropy of the A phase, which indicates that the vortices are continuous, i.e., consist of the A phase even in the core. In finite magnetic fields, a qualitatively similar continuous vortex structure was found, a result compatible with the earlier nuclear magnetic resonance (NMR) experiments, but also a new vortex type with an anomalously high _c was seen when the vortex sample was prepared adiabatically. The observed strong ion focusing indicates broken w-symmetry in the core textures. This is in contradiction with the earlier numerical calculations on the optimal continuous vortex texture. Experiments to observe trapping of ions into the vortex cores were performed both in A and B phases. No trapping could be detected. Mobility measurements in stationary liquid were extended down toT=0.3T _c . Comparison with the calculated transport coefficients yields _A (T)=1.32 _BCS (T) for the maximal A phase gap atP=29.3 bar, and _B (T)=1.12 _BCS (T) for the B phase gap at 4.8 bar. In the normal Fermi liquid a shallow minimum of ion mobility was detected aroundT2T _c =5 mK.     

Metallocene‐based functionalized polyolefins as compatibilizers in polyolefin nanocomposites

This article reports a study of some functionalized polyolefins evaluated as compatibilizers in polyethylene nanocomposites. The functionalized polymers were prepared by direct metallocene‐mediated copolymerizations of ethylene and a functional comonomer. The prepared nanocomposites were evaluated for mechanical and barrier property enhancement. A good combination of mechanical and barrier properties was obtained with the metallocene‐based functionalized polyethylene. The toughness–stiffness balance was better than or comparable to that achieved with conventional functionalized polymers such as maleic anhydride grafted polyethylene. The results also indicated that these metallocene‐based functionalized polyolefins, when used as compatibilizers, could have relatively higher molar masses and lower functionality than those of conventional post‐reactor‐modified compatibilizers, and so the drawbacks associated with the latter could be avoided. Their inherent properties could also further improve the final nanocomposite properties. This was attributed to the more homogeneous nature of metallocene‐catalyzed polymers in comparison with post‐reactor‐modified products. © 2004 Wiley Periodicals, Inc. J Appl Polym Sci 94: 1094–1100, 2004     

Bridging the physical and virtual worlds by local connectivity-based physical selection

The prevalent visions of ambient intelligence emphasise natural interaction between user and functions and services embedded in the environment or available through mobile devices. In these scenarios the physical and virtual worlds seamlessly gear into each other, making crossing the border between these worlds natural or even invisible to the user. The bottleneck in reaching these scenarios appear in the natural mapping between the physical objects and their virtual counterparts. The emergence of local connectivity in mobile devices opens possibilities for implementing novel user interface paradigms to enhance this mapping. We present physical selection paradigm for implementing an intuitive human technology interaction for mobile devices. In order to demonstrate the feasibility of the paradigm we implemented two experimental set-ups using commercially available smart phones with IrDA connectivity. The experiments involved selecting a website by physically pointing at its symbol and making a phone call by pointing at an icon representing the person to be called. In tentative user experiments the physical selection method was more time-efficient and it was perceived more positively by the users than a conventional method.

Concerted utility review of VVER-440 safety research needs (VERSAFE)

There are twelve operating VVER-440/213 reactors in the Czech Republic, Hungary and Slovakia. The plant owners have carried out extensive safety enhancement programmes, which have included measures to improve prevention, mitigation and control of design basis accidents. Recently, there have been a number of projects to study the Severe Accident Management aspects of these plants. The plant owners have also initiated Plant Life Management programmes. The objective of the VERSAFE Concerted Action is to create a network of the VVER-440/213 plant owners and operators, which aims at definition of the further research needs of severe accidents and plant ageing from the utilities’ viewpoint. The starting point is to formulate a general approach to severe accident management and plant life management and then proceed to define the needs for the additional information expected from the safety research. The role of national research institutes and organisations of the partner countries is of crucial importance in performing such research and, thus, creating and maintaining the expertise on the national level. The successful completion of the project will assist in establishing well-defined research projects that are oriented to the needs of the end-users and provide information that can be used to facilitate and support the negotiations of the EU applicant countries operating VVER-440 reactors.     

Mannans as stabilizers of oil-in-water beverage emulsions

The stabilizing effect of spruce galactoglucomannan (GGM) on a model beverage emulsion system was studied and compared to that of guar gum and locust bean gum galactomannans, konjac glucomannan, and corn arabinoxylan. In addition, guar gum was enzymatically modified in order to examine the effect of the degree of polymerization and the degree of substitution of galactomannans on emulsion stability. Use of GGM increased the turbidity of emulsions both immediately after preparation and after storage of up to 14 days at room temperature. GGM emulsions had higher turbidity than the emulsions containing other mannans. The initial turbidity increased with increasing GGM content, but after 14 days storage at room temperature, the turbidity was the highest for GGM/oil ratio of 0.10:1 when ethanol-precipitated GGM was used. Increasing the storage temperature to +45 °C led to rapid emulsion breakdown, but a decrease in storage temperature increased emulsion stability after 14 days. Confocal microscopy showed that the average particle size in the bottom part of GGM emulsions stored for 14 days was smaller than 1 μm. A low degree of polymerization and a high degree of substitution of the modified galactomannans were associated with a decrease in emulsion turbidity.     

Angiotensin I-converting enzyme inhibitory properties of whey protein digests: concentration and characterization of active peptides

The aim of this study was to identify whey-derived peptides with angiotensin I-converting enzyme (ACE) inhibitory activity. The bovine whey proteins alpha-lactalbumin and beta-lactoglobulin were hydrolysed with pepsin, trypsin, chymotrypsin, pancreatin, elastase or carboxypeptidase alone and in combination. The total hydrolysates were fractionated in a two step ultrafiltration process, first with a 30 kDa membrane and then with a 1 kDa membrane. Inhibition of ACE was analysed spectrophotometrically. The peptides were isolated by chromatography and identified by mass and sequencing analysis. The most potent inhibitory peptides were synthesized by the 9-fluorenylmethoxycarbonyl solid phase method. Inhibition of ACE was observed after hydrolysis with trypsin alone, and with an enzyme combination containing pepsin, trypsin and chymotrypsin. Whey protein digests gave a 50% inhibition (IC50) of ACE activity at concentration ranges within 345-1733 micrograms/ml. The IC50 values for the 1-30 kDa fractions ranged from 485 to 1134 micrograms/ml and for the < 1 kDa fraction from 109 to 837 mg/ml. Several ACE-inhibitory peptides were isolated from the hydrolysates by reversed-phase chromatography, and the potencies of the purified peptide fractions had IC50 values of 77-1062 microM. The ACE-inhibitory peptides identified were alpha-lactalbumin fractions (50-52), (99-108) and (104-108) and beta-lactoglobulin fractions (22-25), (32-40), (81-83), (94-100), (106-111) and (142-146).     

Development and application of an automated subcatchment generator for SWMM using open data

An open source subcatchment generator program was developed for the Stormwater Management Model (SWMM) to automate tedious stages in the model construction process. The generator divides the investigated area into subcatchments using a uniform computation grid and connects the grid cells together and to the underlying stormwater network. The system was tested by applying it to two small urban catchments with different fractions of impervious surfaces in Helsinki, Finland, using mostly openly available data. The simulated discharge results were compared to measured data and to results obtained from manually built models. The proposed system significantly accelerated the setup of a SWMM modelling project, as the routing between the subcatchments as well as the subcatchment slopes and flow widths were directly derived from the computation grid. Automatically generated and manually constructed SWMM models produced discharge results that differed only slightly from each other.     

Safety assessment of Lactobacillus strains: presence of putative risk factors in faecal, blood and probiotic isolates

The widespread use of immunosuppressive therapy and antimicrobial agents as well as novel probiotics without a long history of safe use has increased requirements for safety assessment of probiotic lactobacilli. In this study 44 faecal, 52 blood and 15 probiotic isolates (including 3 dairy strains) of Lactobacillus were assayed for their adhesion properties to extracellular matrix proteins and mucus, hemolysis, ability to avoid the induction of respiratory burst in peripheral blood mononucleocytes (PMN) and resistance to human serum. Among tested strains adhesion to collagen, fibrinogen and mucus was isolate-specific and no statistically significant differences were obtained between faecal, blood and probiotic isolates. However, blood isolates showed a trend for higher adhesion to mucus than probiotic strains (P=0.07). Probiotic strains induced lower respiratory burst in PMN when compared to the blood isolates (P<0.05). Moreover, there was a positive correlation between adhesion to collagen and induction of respiratory burst for faecal isolates (P<0.05). In the determination of serum resistance, probiotic strains showed a trend for lower sensitivity to human serum-mediated killing when compared to the faecal isolates (P=0.07). None of the measurable virulence factors were found to be present at statistically higher level in clinical blood isolates when compared to faecal and/or probiotic isolates indicating that these factors do not cause risk when safety of probiotics is considered. However, the significance of adhesion to mucus, low induction of respiratory burst in PMN and resistance to human serum-mediated killing may need further evaluation in experimental animal models and in epidemiological data.