Activation of latent efferent sympathetic fibers in a viscero-visceral reflex circle under conditions of visceral pain in rats

Activation of “silent” efferent fibers due to stimulation of the mesenteric nerve within a definite frequency range is described; the effect is supposed to result from sensitization in reflex circles related to visceral pain. Neirofiziologiya/Neurophysiology, Vol. 38, No. 4, pp. 368–369, July–August, 2006.     

Printed glycan array: antibodies as probed in undiluted serum and effects of dilution

Using printed glycan array (PGA) we compared the results of antibody profiling in undiluted, moderately (1:15) and highly (1:100) diluted human blood serum. Undiluted serum is suitable for studying blood as a tissue in its native state, whereas to study the serum of newborns or small animals one usually has to dilute the starting material in order to have sufficient volume for PGA experimentation. The PGA used in this study allows for the use of whole serum without modifications to the protocol, and the background is surprisingly low. Antibodies profiles observed in undiluted serum versus 1:15 dilution were similar, with only a limited number of new signals identified in the undiluted serum. However, unexpected irregularities were found when IgG and IgM are measured separately, namely, at a 1:15 dilution more intensive IgG signals for many glycans are observed. We believe that in conditions of moderate dilution IgG and IgM antibodies can compete with each other for antigen and as a result, the higher affinity anti-glycan IgGs give rise to more intense signals. Therefore depending on the purpose, different dilutions of serum will be optimal: in competitive 1:15 conditions the observed IgG/IgM ratio corresponds to their titer, whereas at 1:100 dilution the measured ratio corresponds to real molar concentration of IgG and IgM.     

New insights into cartilage repair - the role of migratory progenitor cells in osteoarthritis

Osteoarthritis is one of the most common musculo-skeletal diseases with a complex patholoy and a strong impact on cell biology, differentiation and migration behavior of mesenchymal stem cell-derived progenitor cells. In this review, we elucidate the influence of the pathologically altered extracellular matrix on progenitor cell behavior. Moreover, we discuss the modulation of progenitor cells especially of previously characterized chondrogenic progenitor cells (Koelling et al., 2009) in situ to enhance their regeneration potential. These options comprise the application of growth factors like fibroblast growth factor-2, a Runx-2 knock down and a contemporary anti-inflammatory therapy. This supports endogenous regeneration on behalf of the diseased osteoarthritic cartilage, which otherwise results mainly in an insufficient fibro-cartilaginous repair tissue. Furthermore, new results indicate a role of pericytes in osteoarthritis for these repair attempts. We discuss the biological mechanisms potentially leading to new therapeutic options in osteoarthritis to enhance regeneration in situ.     

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.     

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.     

2-Phenyl-9H-purine-6-carbonitrile derivatives as selective cathepsin S inhibitors

Starting from previously disclosed equally potent cathepsin K and S inhibitor 4-propyl-6-(3-trifluoromethylphenyl)pyrimidine-2-carbonitrile 1, a novel 2-phenyl-9H-purine-6-carbonitrile scaffold was identified to provide potent and selective cathepsin S inhibitors.