Application of sterilised human amnion for reconstruction of the ocular surface

BACKGROUND: Despite improved surgical techniques and the use of new medication, healing of corneal and conjunctival defects cannot always be achieved. In this connection the clinical use of human amnion, produced by different techniques, has represented a successful alternative for the past ten years. The purpose of the present investigation was the development of a clinically secure, therapeutically efficient, and easy-to-handle (transport, storage, application) allogenic amnion transplant. PATIENTS AND METHODS: A new method for an amnion preparation, which contains a sterilisation process in peracetic acid and a drying process in a laminar flow cabinet, was developed as an alternative to previous techniques described in the literature. Amnion transplantation was used to treat 41 patients, 36 of them with corneal ulcer. Further indications for amnion transplantation were symblepharon, descemetocele, as well as dehiscence of conjunctiva after cerclage. RESULTS: Seventy-three percent of cases showed postoperative improvement evidenced by constant vision, while 15 percent showed decreased vision. CONCLUSIONS: The study confirms the observations of previous investigators who consider amnion transplantation an efficient therapeutic method for a multitude of eye diseases. The new method described in this report, guarantees patients' safety by using a validated new sterilisation process against infections that can be transmitted by human tissue. At present this method constitutes the only process available in Germany, and is approved by the Federal Institute for Drugs and Medical Products (BfArM) for the manufacture of human amnion transplants as a finished medical product.     

Substrate and product inhibition of hydrogen production by the extreme thermophile,Caldicellulosiruptor saccharolyticus

Substrate and product inhibition of hydrogen production during sucrose fermentation by the extremely thermophilic bacterium Caldicellulosiruptor saccharolyticus was studied. The inhibition kinetics were analyzed with a noncompetitive, nonlinear inhibition model. Hydrogen was the most severe inhibitor when allowed to accumulate in the culture. Concentrations of 5-10 mM H(2) in the gas phase (identical with partial hydrogen pressure (pH(2)) of (1-2) x 10(4) Pa) initiated a metabolic shift to lactate formation. The extent of inhibition by hydrogen was dependent on the density of the culture. The highest tolerance for hydrogen was found at low volumetric hydrogen production rates, as occurred in cultures with low cell densities. Under those conditions the critical hydrogen concentration in the gas phase was 27.7 mM H(2) (identical with pH(2) of 5.6 x 10(4) Pa); above this value hydrogen production ceased completely. With an efficient removal of hydrogen sucrose fermentation was mainly inhibited by sodium acetate. The critical concentrations of sucrose and acetate, at which growth and hydrogen production was completely inhibited (at neutral pH and 70 degrees C), were 292 and 365 mM, respectively. Inorganic salts, such as sodium chloride, mimicked the effect of sodium acetate, implying that ionic strength was responsible for inhibition. Undissociated acetate did not contribute to inhibition of cultures at neutral or slightly acidic pH. Exposure of exponentially growing cultures to concentrations of sodium acetate or sodium chloride higher than ca. 175 mM caused cell lysis, probably due to activation of autolysins.     

Synthesis of 3-tert-butylcatechol by an engineered monooxygenase

Recombinant Escherichia coli JM101 was used for the in vivo biocatalytic synthesis of 3-tert-butyl- catechol. The bacterial strain synthesized the laboratory-evolved variant HbpA(T2) of 2-hydroxybiphenyl 3-monooxygenase (HbpA, EC 1.14.13.44) from Pseudomonas azelaica HBP1. The mutant enzyme HbpA(T2) is able to hydroxylate 2-tert-butylphenol to the corresponding catechol, a reaction that is not catalyzed by the wild-type enzyme. The biotransformation was performed in a 3-L bioreactor for 24 h. To mitigate the toxicity of the 2-tert-butylphenol starting material, we applied a limited substrate feed. Continuous in situ product removal with the hydrophobic resin Amberlite XAD-4 was used to separate the product from culture broth. In addition, binding to the resin stabilized the product, which was important because 3-tert-butylcatechol is very labile in aqueous solution. The productivity of the process was 63 mg L(-1) h(-1) so that after 24 h, 3.0 g of 3-tert-butylcatechol were isolated. Down-stream processing consisted of two steps. First, bound 2-tert-butylphenol and 3-tert-butylcatechol were eluted from Amberlite XAD-4 with methanol. Second, the two compounds were separated over neutral aluminum oxide, which selectively binds the produced catechol but not the phenol substrate. The final purity of 3-tert-butylcatechol was greater than 98%.     

Parasites are GO

The Malaria Genome Sequencing Consortium Meeting was held on 5–6 June 2001 in Cambridge, UK, and was followed by a workshop discussing the generation of new parasite-specific gene ontology (GO) terms. Present at the meeting, which focused primarily on malaria, were representatives from the research community, the sequencing centres, including the Sanger Centre (Cambridge, UK) and The Institute for Genome Research (TIGR; Rockville, MD, USA), the European Bioinformatics Institute (EBI; Cambridge, UK), Plasmo DB and the GO consortium (Cambridge, UK).     

The effect of ajmalicine spiking and resin addition timing on the production of indole alkaloids from Catharanthus roseus cell cultures

The potential for the feedback inhibition of indole alkaloid synthesis was investigated by spiking suspension cultures of Catharanthus roseus with 0, 9, or 18 mg/L ajmalicine on day 0. The production of ajmalicine, catharanthine, and serpentine were inhibited in a dose-dependent manner. The inhibition was transient as the exogenous ajmalicine was ultimately either metabolized in the medium or within the cell. The addition of neutral resin has previously been shown to enhance ajmalicine production. To minimize product inhibition and product metabolism, Amberlite XAD-7 resin was added to immobilized cultures of C. roseus starting on either day 0, 5, or 15, and fresh resin was exchanged for spent resin every 5 days. The addition of resin did not decrease the viability of the culture. Growth was reduced only in cultures with resin added on day 0. Alkaloid production was enhanced to different extents by the timing of resin addition, suggesting that feedback inhibition or product metabolism was present throughout the culture period. Ajmalicine recovery was nearly 100% when the resin was added initially either on day 0 or day 5. Ajmalicine recovery was reduced to 55% when the resin was added later in the culture period starting on day 15, presumably because of resin saturation or the inaccessibility of alkaloids trapped in the vacuole. Delaying the addition of XAD-7 resin until 5 days after the start of the culture resulted in the highest improvement in ajmalicine production, i.e approximately 70% and also resulted in the complete recovery of ajmalicine from the cell.     

A laboratory evaluation of a regulated airflow through wheat at four combinations of temperature and humidity on the productivity of three species of stored product mites

Aeration is a promising alternative to the use of pesticides for the control of storage insects by cooling bulk grain, but its effectiveness against mite pests is neither fully understood nor optimised. For this reason, the productivity of three species of storage mites, Acarus siro, Lepidoglyphus destructor and Tyrophagus longior, was studied in a laboratory-based experiment at four combinations of temperature and humidity (10°C and 70% RH, 10°C and 80% RH, 20°C and 70% RH, 20°C and 80% RH) with and without an airflow (at 10 m³/h/tonne, equalling 2.5 l/s/tonne, in tubes containing 15 g of grain). This is the first time that a study has examined the three principal components of aeration separately from each other. The effect of these factors was different for each species. For A. siro, temperature was the most important factor, while airflow and humidity were of similar but lesser importance. For T. longior, temperature was more important than humidity, while the reverse was true for L. destructor. For these two species, airflow was the least important factor. The airflow decreased the productivity of L. destructor and T. longior but increased the productivity of A. siro. This increase in productivity confirms that, in practice, prevention of mite infestations, in particular A. siro, will require storage of grain at low temperature, relative humidity and moisture content. This revised version was published online in July 2006 with corrections to the Cover Date.     

Pseudopterosin biosynthesis—pathway elucidation,enzymology, and a proposed production method for anti-inflammatory metabolites from<Emphasis Type="Italic"> Pseudopterogorgia elisabethae</Emphasis>

The pseudopterosins are a family of diterpene pentosides isolated from the marine octocoral, Pseudopterogorgia elisabethae. These compounds possess non-steroidal anti-inflammatory and analgesic properties which have been shown to be greater than the industry standard, indomethacin. In our investigations, we are interested in examining the biosynthesis and enzymology of these compounds for the development of a biotechnological production method. We have isolated the pseudopterosin diterpene cyclase product, elisabethatriene, using a radioactivity-guided isolation. This has provided us with an assay to isolate the diterpene cyclase enzyme. The amino acid sequence of the purified diterpene cyclase will facilitate cloning and expression of the gene in a suitable host. In addition, we have identified over 25 novel diterpenes from one of our collections of P. elisabethae. Several of these compounds appear to be involved in pseudopterosin biosynthesis and are presently being evaluated as potential intermediates. These compounds have also been evaluated for anti-inflammatory activity and some possess greater activity than that of the pseudopterosins. We therefore propose a production method utilizing a combination of recombinant enzyme technology and synthetic methods/biocatalysis in order to produce one or more anti-inflammatory metabolites in P. elisabethae.     

Functional priorities in LCA and design for environment

Aim, Scope and Background  The interest in environmental questions has increased enormously during the last decade. Environmental protection has become an issue of strategic importance within the manufacturing industry and many companies are now working in the field of Design for Environment (DFE). The main purpose of DFE is to create products and services for achieving a sustainable society. Designers are widely believed to have a key role in adapting products to a sustainable society and one of the major instruments in the context of Design for Environment is Life Cycle Assessment (LCA). However, product development creates particular challenges for incorporating environmental issues that combine functional and environmental assessment. A natural and important part of product design is to define and analyse the functions of the product. Consequently, the functional unit in LCA is a core issue in DFE. Most recent research in DFE has focused on how to reduce the environmental impact of products throughout their life-cycle by addressing environmental aspects, while little attention has been given to the functionality of the product. Additionally, early product development phases, so called re-think phases, are considered to have the influence on major changes in products in general. These phases have thus the highest potential for changing products and product systems towards a sustainable development. Main Features  This paper discusses an extended functional representation in design for environment methods to evaluate sustainable design solutions, especially in early (re-think) phases of product design. Based on engineering-design science and several case studies, a concept has been developed describing how functional preferences can be visualised in design for environment and product development. In addition, the functional unit in LCA is discussed. The concept is called Functional Profile (FP) and is additionally exemplified in a case study on radio equipment. Discussion  The new functional characterisation concept helps identify functional priorities in design for environment. The Functional Profile is a structured, systematic and creative concept for identifying the necessary functions of a new product. The FP is envisioned to complement existing design for environment methods, not to replace them. Instead of being a product-development tool or method, the concept is an approach that increases understanding of inter-reactions between functional characteristics of products and their environmental characteristics, which furthermore facilitates trade-off decisions. One of the objectives behind the concept is to highlight the importance of balancing functional requirements and environmental impacts, presenting both the advantages and disadvantages of the product. Outlook  A second paper will be produced to complement the functional-environmental characterisation concept in early product development phase, presenting the environmental characterisation part and illustrating correlations between the functional and environmental sides.     

Excretory-secretory product of Paragonimus westermani newly excysted metacercariae inhibits superoxide production of granulocytes stimulated with IgG

It is well known that the cysteine proteases in excretory-secretory product (ESP) of Paragonimus westermani newly excysted metacercariae (PwNEM) are capable of degrading IgG in vitro. Recent evidence suggests that the IgG-coated surface, such as found on parasites, is one of the most effective physiologic stimuli for granulocyte activation. Therefore, this study was designed to investigate the effect of excretory-secretory product (ESP) of PwNEM on superoxide production of granulocytes stimulated with IgG. The 96-well plates were coated with human IgG (0, 10, 30, 100 micrograms/ml) in the absence or presence of ESP. When granulocytes were incubated in the wells coated with human IgG in the presence of ESP, the level of superoxide production of granulocytes was reduced to about 90% when compared to the cells incubated in the wells coated with IgG alone. This inhibitory effect of the ESP on IgG-induced superoxide production of granulocytes was concentration-dependent. These results suggest that ESP secreted by PwNEM may be important in the control of effector functions of granulocytes stimulated with IgG in human paragonimiasis.