Effects of pretreatment with a xanthine oxidase inhibitor on free radical levels during carotid endarterectomy

Objective: Free radicals contribute to the tissue damage caused by ischaemia-reperfusion. The aim of the present study was to investigate whether preoperative antioxidant therapy (allopurinol) affects free radical levels in cerebral venous blood in connection with surgery for carotid artery stenosis.

Materials and methods: Twenty-five patients were randomised into the study. Thirteen were controls and 12 were pretreated with allopurinol the day before surgery. Before, during and after surgery, blood samples were drawn from the ipsilateral jugular vein. Radical levels were measured using the spin trap technique ex vivo using OXANOH as the spin trap. Multivariate statistics were used with Principal Component Analysis and Partial Least Square regression analysis.

Results: Radical levels increased with diabetes, high leukocyte count, high creatinine and a high degree of contralateral stenosis. Radical levels decreased with high age, blood pressure, collateral circulation as well as operation for left-side carotid artery stenosis. After pretreatment with allopurinol, several of the relationships noted in the control group were eliminated, i.e. leukocyte count, side of operation, Betapred pretreatment and collateral circulation.

Conclusions: Radical levels can be determined in connection with surgery for carotid artery stenosis using an ex vivo spin trap method. With preoperative antioxidant therapy the relationships between enhanced radical levels and clinical data, as seen in control subjects, disappeared. This might indicate a beneficial effect of preoperative pretreatment with antioxidants.     

The quaternary structure of DNA polymerase epsilon from Saccharomyces cerevisiae

DNA polymerase epsilon (Pol epsilon) from Saccharomyces cerevisiae consists of four subunits (Pol2, Dpb2, Dpb3, and Dpb4) and is essential for chromosomal DNA replication. Biochemical characterizations of Pol epsilon have been cumbersome due to protease sensitivity and the limited amounts of Pol epsilon in cells. We have developed a protocol for overexpression and purification of Pol epsilon from S. cerevisiae. The native four-subunit complex was purified to homogeneity by conventional chromatography. Pol epsilon was characterized biochemically by sedimentation velocity experiments and gel filtration experiments. The stoichiometry of the four subunits was estimated to be 1:1:1:1 from colloidal Coomassie-stained gels. Based on the sedimentation coefficient (11.9 S) and the Stokes radius (74.5 A), a molecular mass for Pol epsilon of 371 kDa was calculated, in good agreement with the calculated molecular mass of 379 kDa for a heterotetramer. Furthermore, analytical equilibrium ultracentrifugation experiments support the proposed heterotetrameric structure of Pol epsilon. Thus, both DNA polymerase delta and Pol epsilon are purified as monomeric complexes, in agreement with accumulating evidence that Pol delta and Pol epsilon are located on opposite strands of the eukaryotic replication fork.     

Modelling mating success of saproxylic beetles in relation to search behaviour, population density and substrate abundance

We compared the efficiency of two mate-finding strategies exploited by representatives of the beetle families Cisidae and Anobiidae (genus Dorcatoma) that live inside fruiting bodies of wood-decaying fungi. In the Cisidae both sexes are attracted to host odour, but no pheromones seem to be present (nonpheromone strategy). In the Dorcatoma species only the females are attracted to host odour, but having found a host they attract males with a sexual pheromone (pheromone strategy). With a simulation model, we compared the efficiency of the two strategies at four densities of trees hosting fungal fruiting bodies and at three relative densities of insects. We found only small differences in efficiency between the two strategies at high relative densities of conspecific individuals, regardless of host tree density. The pheromone strategy was relatively more efficient when the relative density of insects or the density of host trees decreased. Thus, species adopting the nonpheromone strategy are probably more sensitive to habitat fragmentation and more likely to decline and go extinct at low population densities (because of Allee effects) than species using the pheromone strategy. Copyright 2003 Published by Elsevier Science Ltd on behalf of The Association for the Study of Animal Behaviour.      

A susceptibility gene for psoriatic arthritis maps to chromosome 16q: evidence for imprinting

Several genetic loci have been reported for psoriasis, but none has been specifically linked to psoriatic arthritis (PsA), a condition that affects >10% of patients with psoriasis. A genetic component for PsA is suggested by segregation within families and high concordance among identical twins. We performed a linkage scan to map genes contributing to PsA. We identified 178 patients with PsA out of 906 patients who were included in our genetic study of psoriasis. Using a comprehensive genealogy database, we were able to connect 100 of these into 39 families. We genotyped the patients using a framework marker set of 1,000 microsatellite markers, with an average density of 3 cM, and performed multipoint, affected-only, allele-sharing linkage analysis using the Allegro program. On the basis of the initial results, we genotyped more markers for the most prominent loci. A linkage with a LOD score of 2.17 was observed on chromosome 16q. The linkage analysis, conditioned on paternal transmission to affected individuals, gave a LOD score of 4.19, whereas a LOD score of only 1.03 was observed when conditioned for maternal transmission. A suggestive locus on chromosome 16q has previously been implicated in psoriasis. Our data indicate that a gene at this locus may be involved in paternal transmission of PsA.     

Attachment of Neisseria gonorrhoeae to the cellular pilus receptor CD46: identification of domains important for bacterial adherence

Pili of Neisseria gonorrhoeae mediate binding of the bacteria to human host cells. Membrane cofactor protein (MCP or CD46), a human cell-surface protein involved in regulation of complement activation, acts as a cellular pilus receptor. In this work, we examined which domains of CD46 mediate bacterial adherence. The CD46 expression was quantified and characterized in human epithelial cell lines. N. gonorrhoeae showed the highest adherence to ME180 cells, which have BC1 as the dominant phenotype. The BC isoforms of CD46 were expressed in all cell lines tested. The adherence was not enhanced by high expression of other isoforms, showing that the BC domain of CD46 is important in adherence of N. gonorrhoeae to human cells. To characterize the pilus-binding site within the CD46 molecule, a set of CD46–BC1 deletion constructs were transfected into COS-7 cells. Piliated N. gonorrhoeae attached well to CD46–BC1-expressing COS-7 cells. We show that the complement control protein repeat 3 (CCP-3) and the serine–threonine–proline (STP)-rich domain of CD46 are important for efficient adherence to host cells. Further, partial deletion of the cytoplasmic tail of CD46 results in low bacterial binding, indicating that the cytoplasmic tail takes part in the process of establishing a stable interaction between N. gonorrhoeae and host cells.     

Complement deficiency ameliorates collagen-induced arthritis in mice

Collagen-induced arthritis (CIA) is an experimental animal model of human rheumatoid arthritis being characterized by synovitis and progressive destruction of cartilage and bone. CIA is induced by injection of heterologous or homologous collagen type II in a susceptible murine strain. DBA/1J mice deficient of complement factors C3 (C3(-/-)) and factor B (FB(-/-)) were generated to elucidate the role of the complement system in CIA. When immunized with bovine collagen type II emulsified in CFA, control mice developed severe arthritis and high CII-specific IgG Ab titers. In contrast, the C3(-/-) and FB(-/-) were highly resistant to CIA and displayed decreased CII-specific IgG Ab response. A repeated bovine collagen type II exposure 3 wk after the initial immunization led to an increase in the Ab response in all mice and triggered arthritis also in the complement-deficient mice. Although the arthritic score of the C3(-/-) mice was low, the arthritis in FB(-/-) mice ranked intermediate with regard to C3(-/-) and control mice. We conclude that complement activation by both the classical and the alternative pathway plays a deleterious role in CIA.     

Gold-coated capillary based 2,4-dichlorophenoxyacetic acid chemi-luminescent assays: possibilities towards multianalysis

The application of gold-coated glass capillaries for the design of a sensitive chemiluminescent immunoassay for 2,4-dichlorophenoxyacetic acid (2,4-D) is reported. The gold coating on the glass capillaries has been partially characterized and its effect on enhancing the signal intensity has been measured. A simple photo-multiplier tube-based photon detector is used for this purpose. At least three times improvement in the signal intensity is observed compared to uncoated glass capillaries, with a consequent improvement in the sensitivity of detection. Using such gold-coated glass capillaries, 2,4-D in the range 10(-9) to 10(-13) mol/l is detectable at a precision of +/-15% (CV%) and a limit of detection of 10(-15) mol/l is achievable. The possibility of using such gold-coated capillaries with a portable multianalytical set-up for field studies is also demonstrated.     

Efficacy of the biocontrol yeast Pichia anomala during long-term storage of moist feed grain under different oxygen and carbon dioxide regimens

The yeast Pichia anomala inhibits the spoilage mold Penicillium roqueforti in laboratory experiments with high-moisture wheat in malfunctioning airtight storage. The ability of P. anomala to prevent mold growth during 14 months of grain storage was evaluated in outdoor silos with different air permeabilities. Freshly harvested wheat in 160-kg portions was inoculated with 10(2) colony-forming units (cfu) g(-1) P. roqueforti, alone or together with 10(4) cfu g(-1) P. anomala. During the first month P. anomala increased to about 10(6) cfu g(-1) in the treated silos to reach 10(7) cfu g (-1) after 9 months. Naturally occurring P. anomala in the untreated silos increased from 10(2) to about 10(3) cfu g(-1) during the first month and reached the same level as the treated silos after 9 months. Oxygen levels were reduced below the detection limit within 1 day, while carbon dioxide levels increased to 80-90% during the first month. P. roqueforti did not grow in wheat treated with P. anomala, regardless of silo permeability, but had increased to 10(5) cfu g(-1) in the untreated silos after 14 months of storage.     

Temporal and spatial patterns in recruitment and succession of a temperate marine fouling assemblage: a comparison of static panels and boat hulls during the boating season

Protection against biofouling is essential for efficient operation of boats and ships. Restrictions on the use of traditional, toxic antifouling coatings call for new less toxic methods. Future antifouling strategies will likely be based on more specific action against dominant foulers and will require more detailed information about spatial and temporal differences in fouling communities on artificial substrates. In this study, the recruitment and succession of fouling organisms was examined on artificial (PMMA) panels exposed to natural flow speeds on the Skagerrak coast (Sweden). The recruitment of foulers on static panels was then compared to fouling allowed to develop on boat hulls in surveys of new nontoxic coatings. The temporal and spatial variation in recruitment was examined on a monthly interval within the boating season, from May-September. Furthermore, the succession of the fouling community was examined during the same interval. A total of 12 sessile invertebrate species was recorded on the static panels with dominant foulers being the barnacle Balanus improvisus (July-September) and the mussel Mytilus edulis (June-August). The mean abundance during peak settlement on panels after 1 month's deployment was 370 +/- 134 individuals dm-2 for B. improvisus and 340 +/- 415 individuals dm-2 for M. edulis. The succession of foulers on the panels led to a dominance of M. edulis (maximum of 7470 +/- 2830 individuals dm-2) over B. improvisus (maximum of 2295 +/- 680 individuals dm-2). This was in contrast to the fouling development on boat hulls, examined after 4 months exposure in 3 successive years, where B. improvisus was the dominant species (mean abundance 222 +/- 104 and maximum 713 +/- 527 individuals dm-2). Some boats were covered to an extent of almost 100% by B. improvisus with only a few M. edulis (mean abundance 20 +/- 16). The biomass of fouling on boat hulls varied from average dry weights of 1.3 +/- 1 to 13 +/- 5 g dm-2. These results show that extrapolation from static panels (common in evaluation of antifouling performance) to fouling on boat hulls may be problematic.