Purification and characterization of heparan sulfate from human primary osteoblasts

Heparan sulfate (HS) is a linear, highly variable, highly sulfated glycosaminoglycan sugar whose biological activity largely depends on internal sulfated domains that mediate specific binding to an extensive range of proteins. In this study we employed anion exchange chromatography, molecular sieving and enzymatic cleavage on HS fractions purified from three compartments of cultured osteoblasts-soluble conditioned media, cell surface, and extracellular matrix (ECM). We demonstrate that the composition of HS chains purified from the different compartments is structurally non-identical by a number of parameters, and that these differences have significant ramifications for their ligand-binding properties. The HS chains purified of conditioned medium had twice the binding affinity for FGF2 when compared with either cell surface or ECM HS. In contrast, similar binding of BMP2 to the three types of HS was observed. These results suggest that different biological compartments of cultured cells have structurally and functionally distinct HS species that help to modulate the flow of HS-dependent factors between the ECM and the cell surface.

Advanced numerical characterization of mono-crystalline copper with defects

Based on the embedded atom method (EAM) and molecular dynamics (MD) method, the mono-crystalline copper with different defects is investigated through tension and nanoindentation simulation. The single-crystal copper nanowire with surface defects is firstly studied through tension. For validation, the tension simulations for nanowire without defect are carried out under different temperatures and strain rates. The defects on nanowires are then systematically studied in considering different defects orientation distribution. It is found that the Young’s modulus is insensitive of surface defects and centro-plane defects. However, the yield strength and yield point show a significant decrease due to the different defects. Specially, the 〖45〗^° defect in surface and in (200) plane exerts the biggest influence to the yield strength, about 34.20% and 51.45% decrease are observed, respectively. Different defects are observed to serve as a dislocation source and different necking positions of the nanowires during tension are found. During nanoindentation simulation, dislocation is found nucleating below the contact area, but no obvious dislocation is generated around the nano-cavity. Comparing with the perfect substrate during nanoindentation, the substrate with nano-cavities emerged less dislocations, it is supposed that the nano-cavity absorbed part of the indent energy, and less plastic deformation happened in the defected substrate.

Cloning and characterization of a novel human gene related to vascular endothelial growth factor

This paper describes the cloning and characterization of a new member of the vascular endothelial growth factor (VEGF) gene family, which we have designated VRF for VEGF-related-factor. Sequencing of cDNAs from a human fetal brain library and RT-PCR products from normal and tumor tissue cDNA pools indicate two alternatively spliced messages with open reading frames of 621 and 564 bp, respectively. The predicted proteins differ at their carboxyl ends resulting from a shift in the open reading frame. Both isoforms show strong homology to VEGF at their amino termini, but only the shorter isoform maintains homology to VEGF at its carboxyl terminus and conserves all 16 cysteine residues of VEGF165. Similarity comparisons of this isoform revealed overall protein identity of 48% and conservative substitution of 69% with VEGF189. VRF is predicted to contain a signal peptide, suggesting that it may be a secreted factor. The VRF gene maps to the D11S750 locus at chromosome band 11q13, and the protein coding region, spanning approximately 5 kb, is comprised of 8 exons that range in size from 36 to 431 bp. Exons 6 and 7 are contiguous and the two isoforms of VRF arise through alternate splicing of exon 6. VRF appears to be ubiquitously expressed as two transcripts of 2.0 and 5.5 kb; the level of expression is similar among normal and malignant tissues.

On a Syntactic Characterization of Classification with a Mind Change Bound

Most learning paradigms impose a particular syntax on the class of concepts to be learned; the chosen syntax can dramatically affect whether the class is learnable or not. For classification paradigms, where the task is to determine whether the underlying world does or does not have a particular property, how that property is represented has no implication on the power of a classifier that just outputs 1’s or 0’s. But is it possible to give a canonical syntactic representation of the class of concepts that are classifiable according to the particular criteria of a given paradigm? We provide a positive answer to this question for classification in the limit paradigms in a logical setting, with ordinal mind change bounds as a measure of complexity. The syntactic characterization that emerges enables to derive that if a possibly noncomputable classifier can perform the task assigned to it by the paradigm, then a computable classifier can also perform the same task. The syntactic characterization is strongly related to the difference hierarchy over the class of open sets of some topological space; this space is naturally defined from the class of possible worlds and possible data of the learning paradigm.

Bispectral and trispectral characterization of transition to chaos in the Duffing oscillator

Higher-order spectral (bispectral and trispectral) analyses of numerical solutions of the Duffing equation with a cubic stiffness are used to isolate the coupling between the triads and quartets, respectively, of nonlinearly interacting Fourier components of the system. The Duffing oscillator follows a period-doubling intermittency catastrophic route to chaos. For period-doubled limit cycles, higher-order spectra indicate that both quadratic and cubic nonlinear interactions are important to the dynamics. However, when the Duffing oscillator becomes chaotic, global behavior of the cubic nonlinearity becomes dominant and quadratic nonlinear interactions are weak, while cubic interactions remain strong. As the nonlinearity of the system is increased, the number of excited Fourier components increases, eventually leading to broad-band power spectra for chaos. The corresponding higher-order spectra indicate that although some individual nonlinear interactions weaken as nonlinearity increases, the number of nonlinearly interacting Fourier modes increases. Trispectra indicate that the cubic interactions gradually evolve from encompassing a few quartets of Fourier components for period-1 motion to encompassing many quartets for chaos. For chaos, all the components within the energetic part of the power spectrum are cubically (but not quadratically) coupled to each other.

Immunohistochemical detection of survivin in canine lymphoma

Survivin is a member of the family of proteins known as 'inhibitors of apoptosis proteins'. Survivin has a role in cellular decisions concerning division and survival and is frequently expressed in neoplastic cells. The aim of the present study was to investigate immunohistochemically the expression of survivin in normal canine tissues and in canine lymphoma. A representative range of fetal and adult normal tissues as well as biopsy samples from dogs with lymphoma were assembled in tissue arrays. The lymphomas were classified according to the revised Kiel and to the Revised European American Lymphoma - World Health Organization (REAL-WHO) schemes. Polyclonal and monoclonal antisera cross-reactive with canine survivin identified cytoplasmic expression of the molecule in a broad range of normal canine cells. The same reagents demonstrated cytoplasmic labelling of more than 5% of cells in all 83 lymphoma samples tested with polyclonal antiserum and in 67 of 82 (82%) of samples tested with monoclonal antiserum. Survivin was expressed by a wide range of canine lymphoma subtypes, but the expression of this molecule in normal canine tissues must be considered if novel therapies targeting survivin are applied to the management of canine lymphoma. © 2010 Elsevier Ltd.

Phenotypic characterization of osteoarthritic osteocytes from the sclerotic zones : a possible pathological role in subchondral bone sclerosis

Subchondral bone sclerosis is a well-recognised manifestation of osteoarthritis (OA). The osteocyte cell network is now considered to be central to the regulation of bone homeo-stasis; however, it is not known whether the integrity of the osteocyte cell network is altered in OA patients. The aim of this study was to investigate OA osteocyte phenotypic changes and its potential role in OA subchondral bone pathogenesis. The morphological and phenotypic changes of osteocytes in OA samples were investigated by micro-CT, SEM, histology, im-munohistochemistry, TRAP staining, apoptosis assay and real-time PCR studies. We demonstrated that in OA subchondral bone, the osteocyte morphology was altered showing rough and rounded cell body with fewer and disorganized dendrites compared with the os-teocytes in control samples. OA osteocyte also showed dysregulated expression of osteocyte markers, apoptosis, and degradative enzymes, indicating that the phenotypical changes in OA osteocytes were accompanied with OA subchondral bone remodelling (increased osteoblast and osteoclast activity) and increased bone volume with altered mineral content. Significant alteration of osteocytes identified in OA samples indicates a potential regulatory role of osteocytes in subchondral bone remodelling and mineral metabolism during OA pathogene-sis.