Factors influencing optimal seating pressure after spinal cord injury.

STUDY DESIGN: Retrospective, cross-sectional design. OBJECTIVES: To identify factors that predict unsatisfactory seating pressure in spinal cord-injured (SCI) individuals. SETTING: Seating Clinic at the University Hospital, Norway. METHODS: All wheelchair users with traumatic SCI hospitalized between 1 January 2007 and 31 December 2010 were included. Individual assessment by a team was performed. To measure seating pressure, a computerized seating pad with sensing points 40 × 40 cm was used. Primary end points were defined as satisfactory or unsatisfactory seating position based on measured pressure (more or less 100 mm Hg), clinical findings and physical activity level. To explore possible risk factors for high seating pressure, both univariate and multivariate regression analysis were performed. RESULTS: A total of 75 persons with SCI were assessed, 39 (52%) with unsatisfactory result. Statistical analysis revealed that use of manual wheelchair (odds ratio (OR)=6.86, confidence interval (CI) 1.77-26.63) and history of pressure ulcer (OR=8.47, CI 2.46-29.13) significantly increase the risk of unsatisfactory seating pressure. Paraplegia caused significantly higher risk (OR=2.5, CI 0.99-6.34) in the univariate model, probably because the SCI with tetraplegia do prefer electrically powered wheelchairs. CONCLUSIONS: Use of manually driven wheelchairs and persons with previous pressure ulcer are at significant risk of high seating pressure and consequently developing new pressure ulcers. The patients from these subcategories need close follow-up regarding seating position and prevention of pressure ulcers.

Modelling the maintenance of egg polymorphism in avian brood parasites and their hosts.

In avian brood parasitism, egg phenotype plays a key role for both host and parasite reproduction. Several parrotbill species of the genus Paradoxornis are parasitized by the common cuckoo Cuculus canorus, and clear polymorphism in egg phenotype is observed. In this article, we develop a population genetics model in order to identify the key parameters that control the maintenance of egg polymorphism. The model analyses show that egg polymorphism can be maintained either statically as an equilibrium or dynamically with frequency oscillations depending on the sensitivity of the host against unlike eggs and how the parasite targets host nests with specific egg phenotypes. On the basis of the model, we discuss egg polymorphism observed in parrotbills and other host species parasitized by the cuckoo. We suggest the possibility that frequencies of egg phenotypes oscillate and we appeal for monitoring of cuckoo-host interactions over a large spatiotemporal scale.

Isolation by time and habitat and coexistence of distinct host races of the common cuckoo.

Isolation by time occurs when different populations of a single species reproduce at different times and thereby reduce the probability of interbreeding, potentially causing divergent adaptation to timing of reproduction, eventually resulting in ecological species separated by timing of reproduction. We analysed extensive data on timing of reproduction by different host races of the common cuckoo Cuculus canorus that is an obligate brood parasite laying eggs in the nests of many different species of passerine birds. Because different hosts breed at different times, specific host races of cuckoos have adapted to specific hosts by laying eggs when nests of these hosts are available, and such divergence may be further exaggerated by differences in timing of breeding among host races with similar habitat requirements. Host species accounted for a quarter of the variance in timing of breeding by the cuckoo. Common cuckoos reproduced at a similar, but narrower subset of dates as did possible hosts, showing that only a fraction of hosts with specific breeding dates were parasitized. Common cuckoo eggs laid in the 'right' kind of nests, phenotypically matching the eggs of the host, were laid later during the season than cuckoo eggs laid in the 'wrong' kind of nests where the eggs did not mimic those of the host. Pairs of sympatric cuckoo host races differed more in timing of breeding than pairs of allopatric host races, and pairs of cuckoo host races with similar breeding habitat differed more in breeding date than pairs of cuckoo host races with dissimilar habitat, as expected from reproductive character displacement. These findings are consistent with cuckoo host races being isolated by timing of breeding and habitat.

The common cuckoo Cuculus canorus is not locally adapted to its reed warbler Acrocephalus scirpaceus host.

The obligate avian brood parasitic common cuckoo Cuculus canorus comprises different strains of females that specialize on particular host species by laying eggs of a constant type that often mimics those of the host. Whether cuckoos are locally adapted for mimicking populations of the hosts on which they are specialized has never been investigated. In this study, we first explored the possibility of local adaptation in cuckoo egg mimicry over a geographical mosaic of selection exerted by one of its main European hosts, the reed warbler Acrocephalus scirpaceus. Secondly, we investigated whether cuckoos inhabiting reed warbler populations with a broad number of alternative suitable hosts at hand were less locally adapted. Cuckoo eggs showed different degrees of mimicry to different reed warbler populations. However, cuckoo eggs did not match the egg phenotypes of their local host population better than eggs of other host populations, indicating that cuckoos were not locally adapted for mimicry on reed warblers. Interestingly, cuckoos exploiting reed warblers in populations with a relatively larger number of co-occurring cuckoo gentes showed lower than average levels of local adaptation in egg volume. Our results suggest that cuckoo local adaptation might be prevented when different cuckoo populations exploit more or fewer different host species, with gene flow or frequent host switches breaking down local adaptation where many host races co-occur.

Rapid change in host use of the common cuckoo Cuculus canorus linked to climate change.

Parasites require synchrony with their hosts so if host timing changes with climate change, some parasites may decline and eventually go extinct. Residents and short-distance migrant hosts of the brood parasitic common cuckoo, Cuculus canorus, have advanced their phenology in response to climate change more than long-distance migrants, including the cuckoo itself. Because different parts of Europe show different degrees of climate change, we predicted that use of residents or short-distance migrants as hosts should have declined in areas with greater increase in spring temperature. Comparing relative frequency of parasitism of the two host categories in 23 European countries before and after 1990, when spring temperatures in many areas had started to increase, we found that relative parasitism of residents and short-distance migrants decreased. This change in host use was positively related to increase in spring temperature, consistent with the prediction that relative change in phenology for different migrant classes drives host-use patterns. These findings are consistent with the hypothesis that climate change affects the relative abundance of different host races of the common cuckoo.

Egg phenotype differentiation in sympatric cuckoo Cuculus canorus gentes.

The brood parasitic common cuckoo Cuculus canorus consists of gentes, which typically parasitize only a single host species whose eggs they often mimic. Where multiple cuckoo gentes co-exist in sympatry, we may expect variable but generally poorer mimicry because of host switches or inter-gens gene flow via males if these also contribute to egg phenotypes. Here, we investigated egg trait differentiation and mimicry in three cuckoo gentes parasitizing great reed warblers Acrocephalus arundinaceus, marsh warblers Acrocephalus palustris and corn buntings Miliaria calandra breeding in close sympatry in partially overlapping habitat types. The three cuckoo gentes showed a remarkable degree of mimicry to their three host species in some but not all egg features, including egg size, a hitherto largely ignored feature of egg mimicry. Egg phenotype matching for both background and spot colours as well as for egg size has been maintained in close sympatry despite the possibility for gene flow.

Avian colour perception predicts behavioural responses to experimental brood parasitism in chaffinches.

Hosts of cuckoos have evolved defences allowing them to discriminate and reject parasite eggs. Mechanisms of discrimination are mostly visually mediated, and have been studied using approaches that do not account for what the receiver (i.e. host) actually can discriminate. Here, for the first time we apply a perceptual model of colour discrimination to study behavioural responses to natural variation in parasite egg appearance in chaffinches Fringilla coelebs. Discrimination of parasite eggs gradually increased with increasing differences in chromatic contrasts as perceived by birds between parasite and host eggs. These results confirm that colour differences of the eggs as perceived by birds are important integral parts of a matching signal used by chaffinch hosts.

Rapid increase in cuckoo egg matching in a recently parasitized reed warbler population.

Parasitic cuckoos lay eggs that mimic those of their hosts, and such close phenotypic matching may arise from coevolutionary interactions between parasite and host. However, cuckoos may also explicitly choose hosts in a way that increases degree of matching between eggs of cuckoos and parasites, with female preference for specific host phenotypes increasing the degree of matching. We tested for temporal change in degree of matching between eggs of the parasitic European cuckoo (Cuculus canorus) and its reed warbler (Acrocephalus scirpaceus) host during 24 consecutive years in a recently parasitized reed warbler population. Cuckoo-host egg matching in an ultraviolet-brownness component yielding most of the chromatic variance of eggs improved during the study period. Improved matching was not due to changes in cuckoo egg phenotype. Cuckoo eggs matched host eggs for ultraviolet-brownness within nests irrespective of duration of sympatry. Ultraviolet-brownness of cuckoo eggs was similar to that of reed warbler eggs at parasitized nests, but differed from that of reed warbler eggs at unparasitized nests. These findings provide tentative support for the cuckoo preference hypothesis suggesting that cuckoo-host egg matching could partially be due to cuckoo females selecting host nests based on the appearance of their eggs.

Improved chitosan-mediated gene delivery based on easily dissociated chitosan polyplexes of highly defined chitosan oligomers.

Nonviral gene delivery systems based on conventional high-molecular-weight chitosans are efficient after lung administration in vivo, but have poor physical properties such as aggregated shapes, low solubility at neutral pH, high viscosity at concentrations used for in vivo delivery and a slow dissociation and release of plasmid DNA, resulting in a slow onset of action. We therefore developed highly effective nonviral gene delivery systems with improved physical properties from a series of chitosan oligomers, ranging in molecular weight from 1.2 to 10 kDa. First, we established structure-property relationships with regard to polyplex formation and in vivo efficiency after lung administration to mice. In a second step, we isolated chitosan oligomers from a preferred oligomer fraction to obtain fractions, ranging from 10 to 50-mers, of more homogeneous size distributions with polydispersities ranging from 1.01 to 1.09. Polyplexes based on chitosan oligomers dissociated more easily than those of a high-molecular-weight ultrapure chitosan (UPC, approximately a 1000-mer), and released pDNA in the presence of anionic heparin. The more easily dissociated polyplexes mediated a faster onset of action and gave a higher gene expression both in 293 cells in vitro and after lung administration in vivo as compared to the more stable UPC polyplexes. Already 24 h after intratracheal administration, a 120- to 260-fold higher luciferase gene expression was observed compared to UPC in the mouse lung in vivo. The gene expression in the lung was comparable to that of PEI (respective AUCs of 2756+/-710 and 3320+/-871 pg luciferase x days/mg of total lung protein). In conclusion, a major improvement of chitosan-mediated nonviral gene delivery to the lung was obtained by using polyplexes of well-defined chitosan oligomers. Polyplexes of oligomer fractions also had superior physicochemical properties to commonly used high-molecular-weight UPC.

Macromolecular triplex zipping observed in derivatives of fungal (1 --> 3)-beta-D-glucan by electron and atomic force microscopy.

Scleroglucan, a comb-like branched (1 --> 3)-beta-D-glucan, dissolves in water as a stiff, triple-helical structure with the single glucose branches extending from the surface. The aim of this study is to investigate structural changes in the triple-helical structure associated with selective chemical modification of the side chains. Electron and atomic force microscopy, respectively, were used to investigate the macromolecular structures of aldehyde and carboxylated derivatives of scleroglucan-namely, scleraldehyde and sclerox-with different degrees of substitution. Scleraldehyde was observed to have structures resembling the triplex of the unmodified scleroglucan for all degrees of substitution up to 1.0. Additionally, an increasing tendency to aggregate for the higher degrees of substitution was observed. Fully carboxylated scleroglucan, sclerox(1.0), prepared from solutions at ionic strengths below 1.0M, revealed dispersed, flexible, coil-like structures. This indicates an electrostatic-driven strand separation of the scleroglucan triple-helical structure occurring concomitant with an increasing fraction of the side chains bearing carboxylate groups. Annealed sclerox(1.0) samples in aqueous 1.0 and 1.5M NaCl exhibited partly, or completely, reassociated triplex ensembles, with species ranging from apparently fully zipped linear and circular topologies, partly zipped structures with triplex strand separation occurring at the ends, to dispersed single-strands with random coil-like appearance. This study shows that periodate oxidation of the scleroglucan side chains is not a sufficient modification of the side chains to induce dissociation of the triple-helical structure, whereas further oxidation of the side chains to carboxylic groups dissociates the triple-helical structure when the degree of substitution is above 0.6.

The cytokine stimulating activity of (1-->3)-beta-D-glucans is dependent on the triple helix conformation.

The immunomodulating properties of comb-like branched (1-->3)-beta-D-glucans scleroglucan, schizophyllan and lentinan depend on branching pattern, molecular weight and higher-order structure. The effect of weight average molecular weight Mw and higher order structure of scleroglucan, on stimulation of human monocytes cultured in vitro to secrete tumor necrosis factor-alpha (TNF-alpha) was investigated. The higher order structures of the scleroglucan samples were determined by electron microscopy. The data showed that the samples with a linear wormlike, triple helical structure with Mw less than 50 x 10(4) g/mol or larger than 110 x 10(4) g/mol stimulated the monocytes more efficiently than samples with Mw in the range (67-110) x 10(4) g/mol. The denaturation of the linear triple helices by NaOH (> 0.25 M), followed by neutralization yielded blends of linear and macrocyclic topologies with concomitant irreversible reduction of the cytokine inducing activity compared with the untreated scleroglucans. The dose-dependent ability to activate monocytes to cytokine production was not restored following annealing of the denatured-renatured samples, despite the fact that electron micrographs revealed similar structures of these annealed samples to the starting material. Pre-incubation of monocytes with antibodies against cluster of differentiation antigens CD14 or CD11b reduced the scleroglucan potency to stimulate TNF-alpha secretion mainly for mAb against CD14 in the presence of serum.

[Genetic counseling in presymptomatic testing for Huntington disease]. / Genetisk veiledning ved presymptomatisk testing for Huntingtons sykdom.

Norwegian law and international guidelines require genetic counselling before, during and after presymptomatic testing for Huntington's disease. The genetic counselling of at-risk persons who considers taking tests, includes explanation of the possible implications of a test result for both participant and relatives. The test is performed only when explicitly requested by the participant and after informed consent. The participant decides if and when the test should be conducted. The participant also has major influence on the timing of the consecutive phases of the testing procedure, in compliance with medical and ethical recommendations. This paper reviews main issues raised during genetic counselling and the preparation period preceding the test and communication of the test result. We illustrate different individual situations and backgrounds for considering presymptomatic testing for Huntington's disease by describing three anonymized cases and associated pedigrees.

The recombinant Azotobacter vinelandii mannuronan C-5-epimerase AlgE4 epimerizes alginate by a nonrandom attack mechanism.

The Ca2+-dependent mannuronan C-5-epimerase AlgE4 is a representative of a family of Azotobacter vinelandii enzymes catalyzing the polymer level epimerization of beta-D-mannuronic acid (M) to alpha-L-guluronic acid (G) in the commercially important polysaccharide alginate. The reaction product of recombinantly produced AlgE4 is predominantly characterized by an alternating sequence distribution of the M and G residues (MG blocks). AlgE4 was purified after intracellular overexpression in Escherichia coli, and the activity was shown to be optimal at pH values between 6.5 and 7.0, in the presence of 1-3 mM Ca2+, and at temperatures near 37 degrees C. Sr2+ was found to substitute reasonably well for Ca2+ in activation, whereas Zn2+ strongly inhibited the activity. During epimerization of alginate, the fraction of GMG blocks increased linearly as a function of the total fraction of G residues and comparably much faster than that of MMG blocks. These experimental data could not be accounted for by a random attack mechanism, suggesting that the enzyme either slides along the alginate chain during catalysis or recognizes a pre-existing G residue as a preferred substrate in its consecutive attacks.

Determination of enzymatic hydrolysis specificity of partially N-acetylated chitosans.

A new method for determining the specificity of hydrolysis of the linear binary heteropolysaccharide chitosan composed of (1-->4)-linked 2-acetamido-2-deoxy-beta-D-glucopyranose (GlcNAc; A-unit) and 2-amino-2-deoxy-beta-D-glucopyranose (GlcN; D-unit) residues is described. The method is based on the assignments of the 13C chemical shifts of the identity (A- or D-units) of the new reducing and non-reducing ends and the variation in their nearest neighbours, using low molecular weight chitosans with known random distribution of A- and D-units as substrate. A highly N-acetylated chitosan with fraction of acetylated units (FA) of 0.68 and a number-average degree of polymerization (DPn) of 30 was hydrolysed with hen egg-white lysozyme, showing that both the new reducing and non-reducing ends consisted exclusively of A-units, indicating a high specificity for A-units in subsites DL and EL on lysozyme. Our data suggests that the preceding unit of the reducing A-units, is invariable, and based on earlier studies, most probably an A-unit, while the unit following the non-reducing A-units can be either an A- or a D-unit. A more detailed study of the specificity of lysozyme at subsite DL was performed by hydrolyzing a more deacetylated chitosan (FA = 0.35 and DPn of 20) to a DPn of 9, showing that even for this chitosan more than 90% of the new reducing ends were acetylated units. Thus, lysozyme depolymerizes partially N-acetylated chitosans by preferentially hydrolyzing sequences of acetylated units bound to site CL, DL and EL of the active cleft, while there is no specificity between acetylated and deacetylated units to site FL. In addition, a moderately N-acetylated chitosan with fraction of acetylated units (FA) of 0.35 and a DPn of 20 was hydrolysed with Bacillus sp. No. 7-M chitosanase, showing that both the new reducing and non-reducing ends consisted exclusively of D-units. Our data suggests that the nearest neigbour to the D-unit at the reducing end is invariable, and based on earlier studies, most probably a D-unit, while the unit following the non-reducing D-units can be either an A- or a D-unit. We conclude that the Bacillus chitosanase hydrolyzes partially N-acetylated chitosan by preferentially attacking sequences of three consecutive deacetylated units, hypothetical subsites CC, DC and EC, where the cleavage occur between sugar units bound to subsites DC and EC. A hypothetical subsite FC on the chitosanase show no specificity with respect to A- and D-units. The new NMR method described herein offers a time and labour-saving alternative to the procedure of extensive hydrolysis of the binary heteropolysaccharide chitosan and subsequent isolation and characterization of the oligosaccharides.

Conformation of (2-->1)-beta-D-fructan in aqueous solution.

The conformation and dilute solution properties of (2-->1)-beta-D-fructan in aqueous solution were studied by gel permeation chromatography, low-angle laser light-scattering photometry, viscometry, small-angle X-ray scattering and electron microscopy. Fractions covering a broad range of weight-average molecular weights (Mw) from 1.49 x 10(4) to 5.29 x 10(6) were obtained from a native sample by ultrasonic degradation and fractional precipitation. For Mw < 4 x 10(4), the intrinsic viscosity [eta] varies with Mw0.71, indicating that the fructan chain behaves as a random coil expanded by an excluded-volume effect in this molecular weight region. For Mw > 10(5), [eta] exhibits an unusually weak dependence on Mw and finally becomes almost independent of molecular weight. This behaviour is interpreted in terms of a globular conformation of the high-molecular-weight fructan molecules. Small-angle X-ray-scattering measurements and electron microscopic observations support this interpretation of the values of [eta] observed.

An antitumor, branched (1-->3)-beta-D-glucan from a water extract of fruiting bodies of Cryptoporus volvatus.

A water-soluble, (1-->6)-branched (1-->3)-beta-D-glucan (H-3-B) was isolated from a hot-water extract of the fruiting bodies of the fungus, Cryptoporus volvatus (Basidiomycetes). Enzymatic analysis using exo-(1-->3)-beta-D-glucanase and methylation analysis indicated that this polysaccharide has a main chain composed of beta-(1-->3)-linked D-glucopyranosyl residues, and single, beta-(1-->6)-linked D-glucopyranosyl residues attached as side chains to, on average, every fourth sugar residue of the main chain. This structure was confirmed by 13C NMR spectra of the glucan in Me2SO-d6. The weight-average molecular weight (Mw) of H-3-B was determined to be 44.0 x 10(4) by gel permeation chromatography equipped with a low-angle laser-light-scattering photometer. The electron microscopic observations showed that H-3-B and its sonicated sample (S-H-3-B, Mw = 13.7 x 10(4)) can be described as linear worm-like chains. The mass per unit length for native and sonicated H-3-B was determined to be 1750 and 1780 g mol-1 nm-1, respectively, from the contour lengths obtained by electron microscopy and the molecular weights. These values are in good agreement with that expected for the triple stranded structure. A sample denatured in 0.1 M NaOH and subsequently renatured by neutralization showed a mixture of linear and cyclic structures, and larger aggregates with less well-defined morphology. The H-3-B and S-H-3-B had antitumor activity against the Sarcoma 180 tumor.

Conformation, order-disorder conformational transitions and gelation of non-crystalline polysaccharides studied using electron microscopy.

Direct imaging of polysaccharides using transmission electron microscopy (EM) is an important alternative to physical characterization of non-crystalline polysaccharides in solution. The polymer nature of stiff-chain polysaccharides is quite apparent from direct visualization of the electron micrographs, despite the fact that commonly employed preparation techniques reduce the resolution limit to about 1-2 nm. Electron microscopy has recently been used to study polysaccharides with emphasis both on quantitative properties like contour length, end-to-end distance and chain stiffness, and on qualitative structural features such as cyclization at the macromolecular level. The structural richness observed for polysaccharides of the beta-D0glucan family after a denaturation-renaturation treatment of the specimen, in particular, illustrates the unique potential of EM as a tool for obtaining conformational information about carbohydrate macromolecules. Examples of the latter also include the recent discoveries of cyclic beta-D-glucan and l-carrageenan structures. The EM technique provides information that is not only complementary to what can be obtained using other physical techniques, but also offers important insight otherwise masked by the averaging implicit in most physical techniques used to study aqueous polysaccharide solutions.

Physicochemical properties of (1-->6)-branched (1-->3)-beta-D-glucans. 1. Physical dimensions estimated from hydrodynamic and electron microscopic data.

The physical dimensions of several (1-->6) branched (1-->3)-beta-D-glucan samples obtained from different organisms and their derivatives have been studied by electron microscopy, light scattering measurements, viscometry, and gel permeation chromatography. The electron micrographs indicate that in most samples these biopolymers are adequately described as linear worm-like coils. A sample reconstituted from alkaline media appeared as a blend of the linear, circular, and aggregated polymer morphologies. The average mass per unit length, ML = Mw/Lw for the macroscopically linear samples, was estimated to be 2100 +/- 200 g mol-1 nm-1. The parameter ML was determined from the contour lengths obtained by electron microscopy and the molecular weight by light scattering measurements. The observed ML was consistent with the triple-helical structure reported from x-ray diffraction studies and observed degree of side-chain substitution. From the molecular snapshots shown in the electron micrographs, the persistence lengths of these beta-D-glucans were determined to be 140 +/- 30 nm. The experimentally determined intrinsic viscosities were consistent with these estimates of ML and persistence length. Comparison of the molecular weight distributions obtained from gel permeation chromatography and those deduced from the electron micrographs indicates that number and weight average contour lengths are more reliable than z and z + 1 averages.

Macrocyclization of polysaccharides visualized by electron microscopy.

Topological features of the polysaccharides schizophyllan, l-carrageenan and gellan gum were studied using electron microscopy. Electron micrographs of schizophyllan not subjected to any thermal or solvent composition history destabilizing the triple helix, show stiff, linear chains consistent with the structure being triple helical and with contour length proportional to the molecular weight in solution. A blend of linear, cyclic and hairpin topologies and higher molecular weight clusters were observed after renaturation, i.e. return to conditions favouring the triple helical structure, from solvent conditions dissociating the triple helix. Electron micrographs of l-carrageenan in salt-free solution reveal linear extended structures. Addition of 0.15 M LiI to the solution before preparation for electron microscopy, i.e. salt conditions that favour ordering but not gelation, yields a large fraction of cyclic structures with circumference of different lengths. Likewise, adding KCl to aqueous gellan gum changes their appearance from dispersed polymers to suprastrands with several associated chains. Macrocyclic species can also be observed in gellan gum after the addition of a gel-promoting salt. The tendency to form macrocyclic structures in competition with intermolecular aggregates is determined by the three factors: (1) chain stiffness relative to overall length; (2) parallel or antiparallel alignment of interacting chain segments; and (3) polymer concentration. The present study indicates that electron microscopy provides information about the topology adopted by polysaccharides.