Muscle power attenuation by tendon during energy dissipation

An important function of skeletal muscle is deceleration via active muscle fascicle lengthening, which dissipates movement energy. The mechanical interplay between muscle contraction and tendon elasticity is critical when muscles produce energy. However, the role of tendon elasticity during muscular energy dissipation remains unknown. We tested the hypothesis that tendon elasticity functions as a mechanical buffer, preventing high (and probably damaging) velocities and powers during active muscle fascicle lengthening. We directly measured lateral gastrocnemius muscle force and length in wild turkeys during controlled landings requiring rapid energy dissipation. Muscle-tendon unit (MTU) strain was measured via video kinematics, independent of muscle fascicle strain (measured via sonomicrometry). We found that rapid MTU lengthening immediately following impact involved little or no muscle fascicle lengthening. Therefore, joint flexion had to be accommodated by tendon stretch. After the early contact period, muscle fascicles lengthened and absorbed energy. This late lengthening occurred after most of the joint flexion, and was thus mainly driven by tendon recoil. Temporary tendon energy storage led to a significant reduction in muscle fascicle lengthening velocity and the rate of energy absorption. We conclude that tendons function as power attenuators that probably protect muscles against damage from rapid and forceful lengthening during energy dissipation.     

Detection of Pseudomonas aeruginosa in sputum headspace through volatile organic compound analysis

Introduction

Chronic pulmonary infection is the hallmark of Cystic Fibrosis lung disease. Searching for faster and easier screening may lead to faster diagnosis and treatment of Pseudomonas aeruginosa (P. aeruginosa). Our aim was to analyze and build a model to predict the presence of P. aeruginosa in sputa.

Methods

Sputa from 28 bronchiectatic patients were used for bacterial culturing and analysis of volatile compounds by gas chromatography–mass spectrometry. Data analysis and model building were done by Partial Least Squares Regression Discriminant analysis (PLS-DA). Two analysis were performed: one comparing P. aeruginosa positive with negative cultures at study visit (PA model) and one comparing chronic colonization according to the Leeds criteria with P. aeruginosa negative patients (PACC model).

Results

The PA model prediction of P. aeruginosa presence was rather poor, with a high number of false positives and false negatives. On the other hand, the PACC model was stable and explained chronic P. aeruginosa presence for 95% with 4 PLS-DA factors, with a sensitivity of 100%, a positive predictive value of 86% and a negative predictive value of 100%.

Conclusion

Our study shows the potential for building a prediction model for the presence of chronic P. aeruginosa based on volatiles from sputum.     

Use of a library of mutated Maackia amurensis hemagglutinin for profiling the cell lineage and differentiation

Thirty-five variant lectins were prepared by mutations of two amino acids within the carbohydrate-recognition domain of Maackia amurensis hemagglutinin (MAH). Each lectin showed unique carbohydrate specificity according to their bindings to soluble polyacrylamide with various mono- and oligosaccharides and to glycophorin A. The relative intensity of the bindings of carcinoma, myeloid, fibroblastic, and melanoma cells to immobilized MAH variant lectins was examined. Each cell line showed distinct profiles regarding the number of cells bound to wild-type and 35 MAH variants and the differences and the similarities in these binding profiles were quantitatively documented by the cluster analysis. The cell lines were classified into several groups and these groups surprisingly corresponded to the lineage of the cells. These results indicated that a library of mutated MAH is useful as a tool for the profiling of various cells based on the variations of the surface glycans.     

Galectin-loaded cells as a platform for the profiling of lectin specificity by fluorescent neoglycoconjugates: a case study on galectins-1 and -3 and the impact of assay setting

The involvement of galectins as pleiotropic regulators of cell adhesion and growth in disease progression explains the interest to define their ligand-binding properties. Toward this end, it is desirable to approach in vivo conditions to attain medical relevance. In order to simulate physiological conditions with cell surface glycans as recognition sites and galectins as mediators of intercellular contacts we developed an assay using galectin-loaded Raji cells. The extent of surface binding of fluorescent neoglycoconjugates depended on the lectin presence and the type of lectin, the nature of the probes' carbohydrate headgroup and the density of unsubstituted beta-galactosides on the cell surface. Using the most frequently studied galectins-1 and -3, application of this assay led to rather equal binding levels for linear and branched oligomers of N-acetyllactosamine. A clear preference of galectin-3 for alpha1-3-linked galactosylated lactosamine was noted. In parallel, a panel of 24 neoglycoconjugates was tested as inhibitors of galectin binding from solution to N-glycans of surface-immobilized asialofetuin. These two assays differ in presentation of the galectin and ligand, facilitating identification of assay-dependent properties. Under the condition of the cell assay, selectivity among oligosaccharides for the lectins was higher, and extraordinary affinity of galectin-1 to 3'-O-sulfated probes in a solid-phase assay was lost in the cell assay. Having introduced and validated a cell assay, the comprehensive profiling of ligand binding to cell-surface-presented galectins is made possible.     

Structural properties of plant and mammalian lipoxygenases. Temperature-dependent conformational alterations and membrane binding ability

Lipoxygenases form a heterogeneous family of lipid peroxidizing enzymes, which have been implicated in the synthesis of inflammatory mediators, in cell development and in the pathogenesis of various diseases with major health and political relevance (atherosclerosis, osteoporosis). The crystal structures of various lipoxygenase-isoforms have been reported, and X-ray coordinates for enzyme-ligand complexes are also available. Although the 3D-structures of plant and animal lipoxygenase-isoforms are very similar, recent small-angle X-ray scattering data suggested a higher degree of motional flexibility of mammalian isozymes in aqueous solutions. To explore the molecular basis for these differences we performed dynamic fluorescence measurements that allowed us to study temperature-induced conformational changes arising from three-dimensional fluctuations of the protein matrix. For this purpose, we first investigated the impact of elevated temperature on activity, secondary structure, tertiary structure dynamics and conformational alterations. Applying fluorescence resonance energy transfer we also tested the membrane binding properties of the two lipoxygenase-isoforms, and compared their binding parameters. Taken together, our results indicate that the rabbit 12/15-lipoxygenase is more susceptible to temperature-induced structural alterations than the soybean enzyme. Moreover, the rabbit enzyme exhibits a higher degree of conformational flexibility of the entire protein molecule (global flexibility) and offers the possibility of augmented substrate movement at the catalytic center (local flexibility).     

Domain IV of mouse laminin beta1 and beta2 chains.

Domain IV, consisting of about 230 residues, represents a particular protein module so far found only in laminin beta1 and beta2 chains. Both domains were obtained by recombinant production in mammalian cells. They showed a globular structure, as expected from electron microscopic examination of laminins. Fragment beta1IV was obtained as a monomer and a disulfide-bonded dimer, and both were modified to approximately 50% by a single chondroitin sulfate chain attached to Ser721 of an SGD consensus sequence. Dimerization is caused by an odd number of cysteines, with three of them having a partial thiol character. Whether both modifications also occur in tissue forms of laminin remains to be established. Fragment beta2IV was only obtained as a monomer, as it lacked one crucial cysteine and the SGD sequence. It required, however, the presence of two adjacent LE modules for proper folding. Polyclonal antibodies raised against both fragments showed no cross-reaction with each other and allowed establishment of beta chain-specific radioimmunoassays and light and electron microscopic immunostaining of tissues. This demonstrated a 5-25-fold lower content of beta2 compared with beta1 chains in various tissue extracts of adult mice. Tissues derived from beta2-deficient mice failed to react with the beta2-specific antibodies but showed a twofold higher content of beta1 than heterozygotes. The antibodies to beta2 showed broader tissue staining than reported previously, including in particular a distinct reaction with the extrasynaptic endomysium of skeletal muscle. Immunogold staining localized both beta chains primarily to basement membranes of kidney, muscle and various other tissues.     

Synthesis and structure of [bis(pyrazol-1-yl)acetato]trioxorhenium(VII) and [bis(3,5-dimethylpyrazol-1-yl)acetato]trioxorhenium(VII)

The coordination of the ligands bis(pyrazol-1-yl)acetate (bpza) and bis(3,5-dimethylpyrazol-1-yl)acetate (bdmpza) to rhenium(VII) was investigated. The compounds [(bpza)ReO₃] and [(bdmpza)ReO₃] were synthesised by reaction of bpza and bdmpza with perrhenic acid with the loss of one water molecule. The new complex [(bdmpza)ReO₃] was characterised by single-crystal X-ray analysis. It has a monomeric structure with a distorted octahedron for the [N,N,O]ReO₃ central core.