Non-penetrating polymeric cryofixatives for ultrastructural and analytical studies of biological tissues.

Existing freezing methods for biological tissues, either for the purposes of storing living material or for ultrastructural observation, are hampered by various limitations, such as small samples (spray-freezing) or the introduction of physiological and/or cytological alterations (incubation in DMSO or glycerol, high pressure freezing). We have investigated the possibility of using aqueous polymer solutions as extracellular cryofixative media, the basis of structural preservation being the capacity of relatively dilute solutions to vitrify under quench cooling conditions. Evidence is presented to show that two such polymers (polyvinylpyrrolidone and hydroxyethyl starch) control—or even inhibit—intracellular freezing in a wide variety of quench cooled tissue samples. The effects of these polymers on the physiology of tissues from a range of different organisms has been assessed by microscopy, electrophysiology and secretion studies. At the concentrations necessary to ensure vitrification the polymer solutions cause only slight perturbations of the normal functioning of the cells studied. The special application of these studies to freeze fracture and scanning electron microscopy is discussed.     

The application of shear and extensional viscosity measurements to assess the potential of hylan in viscosupplementation

The shear and extensional viscosity characteristics have been compared for hyaluronan and two samples of a cross-linked derivative, hylan, of different molecular weights. While shear thinning behavior was observed for all systems in shear flow, strain thickening was observed in extensional flow for the relatively dilute systems. However, there was a progressive transition to shear thinning behavior as the polymer concentration was increased. It is evident from the results that the shear flow techniques alone provide an incomplete picture of the rheological properties of these materials and that extensional flow characteristics are potentially dominant. For example, at relatively high deformation rates of 500 s¹ and above, our results show that the extensional viscosities of aqueous solutions of the various polymers are at least two orders of magnitude greater than their corresponding shear flow viscosities. The incremental differences in viscosity with concentration increased with increasing molecular mass of the polymers and were greater in exensional flow than shear flow. These results demonstrate that the dynamic network structure formed by the higher molecular mass hylans offer potentially better physical and mechanical properties for viscosupplementation of diseased osteoarthritis joints compared with the parent hyaluronan.     

Adhesion of the marine fouling diatom amphora coffeaeformis to non‐solid gel surfaces

Diatom adhesion to different gel surfaces was tested under different shear conditions, using the fouling marine diatom Amphora coffeaeformis as test organism. Four polymers were selected to obtain a test matrix containing gels with different surface charge as well as different surface energies, viz. agarose, alginate, chitosan and chemically modified polyvinylalcohol (PVA‐SbQ). Three experimental systems were applied to obtain different shear rates. Experimental system 1 consisted of gels cast in a cell culturing well plate for comparing initial adhesion as well as long term biofilm development in the absence of shear. In experimental system 2, microscope slide based test surfaces were tested in aquaria under low shear conditions. A rotating annular biofilm reactor was used to obtain high and controlled shear rates. At high shear rates A. coffeaeformis cells adhered better to the charged polymer gels (alginate and chitosan) than to the low charged polymer gels (agarose and PVA‐SbQ). In the system where shear was absent A. coffeaeformis cells developed a biofilm on agarose equivalent to the charged polymer gels, while adhesion to PVA‐SbQ remained low at all shear rates. It is concluded that non‐solid surfaces did not represent an obstacle to settling and growth of this organism. As observed for solid surfaces, low charge density led to reduced attachment, particularly at high shear.     

Biodegradable co-polymers of amino acids and synthetic polymers

The methods of synthesis and properties of copolymers of natural amino acids and of oligopeptides with synthetic polymers are considered as well as their liability to biodestruction processes both under the effect of proteolytic enzymes in vitro and under conditions of the implantation to animals. Such copolymers may be successfully used in biology and medicine as a material for manufacturing temporal endoprostheses, carriers of medicinal preparations, immobilization of biologically active compounds as well as in studies of the mechanism of their action.     

Cellulose depolymerization to glucose and other water soluble polysaccharides by shear deformation and high pressure treatment

The simultaneous action of shear deformation and high pressure (SDHP) creates changes in the structure of wood and its main components (cellulose, hemicelluloses, lignin). The formation of water and alkali soluble polysaccharides under SDHP action, proceeds in seconds in the solid state, without the use of any reagents and solvents. Therefore, SDHP seems to be a technologically safe method and friendly to the environment. The amorphization of cellulose crystallites and depolymerization of cellulose chains were observed under a wide range of pressures (1–6 GPa), both for cellulose samples and the cellulose part of wood. Similar depolymerization occurs in the hemicellulose part of wood. The decomposition of polysaccharides under SDHP causes the formation of the water soluble part, whose content increases with pressure and the applied shear deformation. A maximum solubility of 40% and 55% was registered at 6 GPa following treatment of cellulose and birch wood samples. A higher output in the case of wood can be explained by a specific role of lignin under SDHP, which acts as a ‘grinding stone’ during cellulose and hemicelluloses destruction. As shown by high-performance size exclusion chromatography, the water soluble fraction obtained from cellulose contained glucose (2.6%), cellobiose (9.6%), cellotriose (16.6%) and other higher water soluble oligomers (71%). Almost complete dissolution (98%) of the treated cellulose sample can be achieved by extraction with 10% NaOH solution. The SDHP treated birch wood was subjected to submerged fermentation (with Trichoderma viride), and a 13% output of proteins was obtained. In this case, the water soluble part played the role of the so called ‘start sugars’. Abbreviations: ASF, alkali soluble fraction; DP, degree of polymerization; EC, energy consumption; HP, high pressure; LMWS, low molecular weight sugars; MC, moisture content; MCC, microcrystalline cellulose; SD, shear deformation, SDHP, shear deformation under high pressure; SS, shear strength; WSF, water soluble fraction This revised version was published online in November 2006 with corrections to the Cover Date.     

Synthesis and characterization of a novel superabsorbent polymer of N,O-carboxymethyl chitosan graft copolymerized with vinyl monomers

A novel superabsorbent polymer was prepared by graft copolymerization of sodium acrylate and 1-vinyl-2-pyrrolidone onto the chain of N,O-carboxymethyl chitosan. The molecular structure of the product was confirmed by FTIR. The surface morphologies before and after the polymerization were examined by SEM. By studying the water absorption of the polymer synthesized under different conditions, the optimal conditions for synthesizing the polymer with the highest swelling ratio was defined. We showed that the water absorption rates of the prepared polymers were high, the swelling processes of the polymers showed first-order kinetics, and the swelling ratio of the polymer was pH dependent.     

Deformation mechanism of leukocyte adhering to vascular surface under steady shear flow

The adhesion of leukocytes to vascular surface is an important biomedical problem and has drawn extensive attention. In this study, we propose a compound drop model to simulate a leukocyte with a nucleus adhering to the surface of blood vessel under steady shear flow. A two-dimensional computational fluid dynamics (CFD) is conducted to determine the local distribution of pressure on the surface of the adherent model cell. By introducing the parameter of deformation index (DI), we investigate the deformation of the leukocyte and its nucleus under controlled conditions. Our numerical results show that: (i) the leukocyte is capable of deformation under external exposed flow field. The deformation index increases with initial contact angle and Reynolds number of external exposed flow. (ii) The nucleus deforms with the cell, and the deformation index of the leukocyte is greater than that of the nucleus. The leukocyte is more deformable while the nucleus is more capable of resisting external shear flow. (iii) The leukocyte and the nucleus are not able to deform infinitely with the increase of Reynolds number because the deformation index reaches a maximum. (iv) Pressure distribution confirms that there exists a region downstream of the cell, which produces high pressure to retard continuous deformation and provide a positive lift force on the cell. Meanwhile, we have measured the deformation of human leukocytes exposed to shear flow by using a flow chamber system. We found that the numerical results are well consistent with those of experiment. We conclude that the nucleus with high viscosity plays a particular role in leukocyte deformation.     

Unimolecular micelles based on hydrophobically derivatized hyperbranched polyglycerols: ligand binding properties

This paper discusses the binding and release properties of hydrophobically modified hyperbranched polyglycerol-polyethylene glycol copolymers that were originally developed as human serum albumin (HSA) substitutes. Their unimolecular micellar nature in aqueous solution has been proven by size measurements and other spectroscopic methods. These polymers aggregate weakly in solution, but the aggregates are broken down by low shear forces or by encapsulating a hydrophobic ligand within the polymer. The small molecule binding properties of these polymers are compared with those of HSA. The preliminary in vitro paclitaxel release studies showed very promising sustained drug release characteristics achieved by these unimolecular micelles.     

Keratin filament suspensions show unique micromechanical properties.

All epithelial cells feature a prominent keratin intermediate filament (IF) network in their cytoplasm. Studies in transgenic mice and in patients with inherited epithelial fragility syndromes showed that a major function of keratin IFs is to provide mechanical support to epithelial cell sheets. Yet the micromechanical properties of keratin IFs themselves remain unknown. We used rheological methods to assess the properties of suspensions of epidermal type I and type II keratin IFs and of vimentin, a type III IF polymer. We find that both types of IFs form gels with properties akin to visco-elastic solids. With increasing deformation they display strain hardening and yield relatively rapidly. Remarkably, both types of gels recover their preshear properties upon cessation of the deformation. Repeated imposition of small deformations gives rise to a progressively stiffer gel for keratin but not vimentin IFs. The visco-elastic moduli of both gels show a weak dependence upon the frequency of the input shear stress and the concentration of the polymer, suggesting that both steric and nonsteric interactions between individual polymers contribute to the observed mechanical properties. In support of this, the length of individual polymers contributes only modestly to the properties of IF gels. Collectively these properties render IFs unique among cytoskeletal polymers and have strong implications for their function in vivo.     

Deformation mechanism of leukocyte adhering to vascular surface under steady shear flow

~~Deformation mechanism of leukocyte adhering to vascular surface under steady shear flow~~     

The relationship between surface tension and the industrial performance of water-soluble polymers prepared from acid hydrolysis lignin, a saccharification by-product from woody materials

In this study, water-soluble anionic and cationic polymers were prepared from sulfuric acid lignin (SAL), an acid hydrolysis lignin, and the relationship between the surface tension of these polymers and industrial performance was examined. The SAL was phenolized (P-SAL) to enhance its solubility and reactivity. Sulfonation and the Mannich reaction with aminocarboxylic acids produced water-soluble anionic polymers and high-dispersibility gypsum paste. The dispersing efficiency increased as the surface tension decreased, suggesting that the fluidity of the gypsum paste increased with the polymer adsorption on the gypsum particle surface. Water-soluble cationic polymers were prepared using the Mannich reaction with dimethylamine. The cationic polymers showed high sizing efficiency under neutral papermaking conditions; the sizing efficiency increased with the surface tension. This suggests that the polymer with high hydrophilicity spread in the water and readily adhered to the pulp surface and the rosin, showing good retention.     

The use of high pressure for separation and production of bioactive molecules

Due to its action on the forces governing inter- and intramolecular interactions, the application of high pressure to biopurification or bio-elaboration of a product are of interest. The two closely thermodynamically related parameters, pressure and temperature, render processes based on their action clean, as no chemical reagents have to be added (and thus further removed) when they are applied. The use of high pressure in the development of desorption methods for the purification of bioactive molecules, particularly in the immunoaffinity field, is reviewed and discussed. Also mentioned is the application of the pressure parameter during the synthesis of a bioreagent. Finally, integrated processes relative to the synthesis and purification of these compounds are proposed.     

Deformation responses of a physically cross-linked high molecular weight elastin-like protein polymer

Recombinant protein polymers were synthesized and examined under various loading conditions to assess the mechanical stability and deformation responses of physically cross-linked, hydrated, protein polymer networks designed as triblock copolymers with central elastomeric and flanking plastic-like blocks. Uniaxial stress-strain properties, creep and stress relaxation behavior, as well as the effect of various mechanical preconditioning protocols on these responses were characterized. Significantly, we demonstrate for the first time that ABA triblock protein copolymers when redesigned with substantially larger endblock segments can withstand significantly greater loads. Furthermore, the presence of three distinct phases of deformation behavior was revealed upon subjecting physically cross-linked protein networks to step and cyclic loading protocols in which the magnitude of the imposed stress was incrementally increased over time. We speculate that these phases correspond to the stretch of polypeptide bonds, the conformational changes of polypeptide chains, and the disruption of physical cross-links. The capacity to select a genetically engineered protein polymer that is suitable for its intended application requires an appreciation of its viscoelastic characteristics and the capacity of both molecular structure and conditioning protocols to influence these properties.     

Osmotic properties of poly(ethylene glycols): quantitative features of brush and bulk scaling laws

From glycosylated cell surfaces to sterically stabilized liposomes, polymers attached to membranes attract biological and therapeutic interest. Can the scaling laws of polymer "brushes" describe the physical properties of these coats? We delineate conditions where the Alexander-de Gennes theory of polymer brushes successfully fits the intermembrane distance versus applied osmotic stress data of Kenworthy et al. for poly(ethylene glycol)-grafted multilamellar liposomes. We establish that the polymer density and size in the brush must be high enough that, in a bulk solution of equivalent monomer density, the polymer osmotic pressure is independent of polymer molecular weight (the des Cloizeaux semidilute regime of bulk polymer solutions). The condition that attached polymers behave as semidilute bulk solutions offers a rigorous criterion for brush scaling-law behavior. There is a deep connection between the behaviors of semidilute polymer solutions in bulk and polymers grafted to a surface at a density such that neighbors pack to form a uniform brush. In this regime, two-parameter unconstrained fits of the Alexander-de Gennes brush scaling laws to the Kenworthy et al. data yield effective monomer lengths of 3.3-3.6 A, which agree with structural predictions. The fitted distances between grafting sites are larger than expected from the nominal mole fraction of poly(ethylene glycol)-lipids; the chains apparently saturate the surface. Osmotic stress measurements can be used to estimate the actual densities of membrane-grafted polymers.     

A finite deformation theory for cartilage and other soft hydrated connective tissues--I. Equilibrium results

The determination of valid stress-strain relations for articular cartilage under finite deformation conditions is a prerequisite for constructing models for synovial joint lubrication. Under physiological conditions of high strain rates and/or high stresses in the joint, large strains occur in cartilage. A finite deformation theory valid for describing cartilage, as well as other soft hydrated connective tissues under large loads, has been developed. This theory is based on the choice of a specific Helmholtz energy function which satisfies the generalized Coleman-Noll (GCN0) condition and the Baker-Ericksen (B-E) inequalities established in finite elasticity theory. In addition, the finite deformation biphasic theory includes the effects of strain-dependent porosity and permeability. These nonlinear effects are essential for properly describing the biomechanical behavior of articular cartilage, even when strain rates are low and strains are infinitesimal. The finite deformation theory describes the large strain behavior of cartilage observed in one-dimensional confined compression experiments at equilibrium, and it reduces to the linear biphasic theory under infinitesimal strain and slow strain rate conditions. Using this theory, we have determined the material coefficients of both human and bovine articular cartilages under large strain conditions at equilibrium. The theory compares very well with experimental results.     

Mini-review: combinatorial approaches for the design of novel coating systems

Combinatorial and high throughput experimental methods are being applied to the design and development of novel polymers and coatings used in a number of application areas. Methods have been developed for polymer synthesis and screening and for the development of polymer thin film and coating libraries and the screening of these libraries for key properties such as surface energy and modulus. Combinatorial and high throughput methods enable the efficient exploration of a large number of compositional variables over a wide range. In the development of coatings for use in the marine environment, the key challenge is in the development of screening methods that can predict good performance. A number of assays are under development that will permit the rapid screening of the interaction of coatings with representative marine organisms.     

Mini-review: Combinatorial approaches for the design of novel coating systems

Abstract

Combinatorial and high throughput experimental methods are being applied to the design and development of novel polymers and coatings used in a number of application areas. Methods have been developed for polymer synthesis and screening and for the development of polymer thin film and coating libraries and the screening of these libraries for key properties such as surface energy and modulus. Combinatorial and high throughput methods enable the efficient exploration of a large number of compositional variables over a wide range. In the development of coatings for use in the marine environment, the key challenge is in the development of screening methods that can predict good performance. A number of assays are under development that will permit the rapid screening of the interaction of coatings with representative marine organisms.     

Regioselectivity of ferulic acid polymerization catalyzed by oxidases

Enzymatic oxidation of ferulic acid catalyzed by oxidases (laccase and peroxidase) was carried out. Ferulic acid was shown to be subjected to oxidative processes leading to the formation of oligomeric and polymeric structures. The polymer formation takes place due to the formation of CAr-CAr-, and CAr-O-CAr-bonds, as well as due to reactions of opening of the propane chain double bond. Different dynamic conditions of the enzymatic reactions were used to study the effects of conditions on the biosynthesis in vitro of some dehydropolymers: the method of dropwise mixing (endwise polymers) and a single addition (bulk polymers). The chemical structures of the resulting compounds were examined by the methods of IR, 1H NMR, and 13N NMR spectroscopy. Differences in the quantitative ratio of structural fragments in a polymer cause changes in its thermal characteristics.     

Biorheological views of endothelial cell responses to mechanical stimuli

Vascular endothelial cells are located at the innermost layer of the blood vessel wall and are always exposed to three different mechanical forces: shear stress due to blood flow, hydrostatic pressure due to blood pressure and cyclic stretch due to vessel deformation. It is well known that endothelial cells respond to these mechanical forces and change their shapes, cytoskeletal structures and functions. In this review, we would like to mainly focus on the effects of shear stress and hydrostatic pressure on endothelial cell morphology. After applying fluid shear stress, cultured endothelial cells show marked elongation and orientation in the flow direction. In addition, thick stress fibers of actin filaments appear and align along the cell long axis. Thus, endothelial cell morphology is closely related to the cytoskeletal structure. Further, the dynamic course of the morphological changes is shown and the related events such as changes in mechanical stiffness and functions are also summarized. When endothelial cells were exposed to hydrostatic pressure, they exhibited a marked elongation and orientation in a random direction, together with development of centrally located, thick stress fibers. Pressured endothelial cells also exhibited a multilayered structure with less expression of VE-cadherin unlike under control conditions. Simultaneous loading of hydrostatic pressure and shear stress inhibited endothelial cell multilayering and induced elongation and orientation of endothelial cells with well-developed VE-cadherin in a monolayer, which suggests that for a better understanding of vascular endothelial cell responses one has to take into consideration the combination of the different mechanical forces such as exist under in vivo mechanical conditions.