Experimental and Monte Carlo simulation studies of the thermodynamics of polyethyleneglycol chains grafted to lipid bilayers

Experimental measurements of the affinity of binding of fluorescent acylated polyethyleneglycol (PEG) conjugates to bilayers containing varying levels of phosphatidylethanolamine-PEGs (PE-PEGs) have been combined with Monte Carlo simulations to investigate the properties of the polymer chains at a PEG-grafted lipid interface. The affinity of binding of such conjugates to large unilamellar phosphatidylcholine/phosphatidylethanolamine (9:1) vesicles decreases 27-fold as the size of the coupled PEG chain increases from 1 to 114 monomer units. Incorporation of increasing amounts of PE-PEG2000 or PE-PEG5000 into the vesicles progressively reduces the affinity of binding of acylpeptide-PEG2000 or -PEG5000 conjugates. Monte Carlo simulations of surfaces with grafted PEG chains revealed no significant dependence of several characteristic properties of the polymer chains, including the average internal energy per polymer and the radii of gyration, on the grafting density in the range examined experimentally. The average conformation of a surface-grafted PEG2000 or PEG5000 chain was calculated to be fairly extended even at low grafting densities, and the projected cross-sectional areas of the grafted PEG chains are considerably smaller than those predicted on the basis of the estimated Flory radius. The experimental variation of the binding affinity of acylated conjugates for bilayers containing varying mole fractions of PE-PEG2000 or -PEG5000 is well explained by expressions treating the surface-grafted PEG polymers either as a van der Waals gas or as a system of rigid discs described by scaled particle theory. From the combined results of our experimental and simulation studies we conclude that the grafted PEG chains exist in a "mushroom" regime throughout the range of polymer densities examined experimentally and that the diminished affinity of binding of acylated-PEG conjugates to bilayers containing PE-PEGs results from occlusion of the surface area accessible for conjugate binding by the mobile PE-PEG polymer chains.     

Grafted poly-(ethylene glycol) on lipid surfaces inhibits protein adsorption and cell adhesion

Monolayers of dipalmitoyl-phosphatidylethanolamine (DPPE) mixing with various mole percentages of distearoyl-phosphatidylethanolamine (DSPE)-conjugated poly-(ethylene glycol) (PEG m.w. 750-5000) were deposited on DPPE-coated glass surfaces by the Langmuir-Blodgett method. Increasing percentages of grafted PEG in these supported lipid surfaces increasingly inhibit the adsorption of bovine serum albumin (BSA), laminin, and fibronectin. Increasing percentages of grafted PEG also inhibit the adhesion of erythrocytes, lymphocytes, and macrophages to these supported lipid surfaces. The adsorption of proteins on lipid coated glass surfaces were assayed by the fluorescence of FITC-labelled proteins. Cell adhesion was measured mainly by microscopic counting. The concentration of PEG-grafted lipids required for the inhibition of erythrocyte adhesion decreases with increasing molecular weight of the grafted PEG. The inhibitory effects are strongly dependent on the graft density of PEG at low concentrations, but weakly dependent on graft density at higher concentrations. For DSPE-PEG5000, the change of graft density dependency occurs approximately at the complete coverage of the lipid surface by the grafted polymer in the mushroom conformation (0.7 mol%), and the transition to partial brush conformation. The change-overs become less distinctive for grafted PEG of lower molecular weights, probably due to the failure of strictly mushroom and brush models of the polymer. The relative inhibitory efficiency is protein or cell dependent. The implication on the function of stealth liposomes is discussed.     

Optimisation of eco-friendly binary binder system for powder injection moulding

Abstract

In recent years, many efforts have been made to obtain more environmentally acceptable powder injection moulding processes. In this sense, the purpose of this study is to optimise an eco-binder based on polyethylene glycol (PEG) as a water soluble component and cellulose acetate butyrate (CAB) as a natural backbone polymer derived from cellulose for powder injection moulding of zirconium silicate powders until a solvent debinding stage. Four different feedstocks have been investigated. As well as, a volume fraction of PEG and CAB 70/30 (vol.-%) and a solid loading of 57·5 (vol.-%) were maintained, molecular weights of polymers were combined in order to minimize distortion during binder solvent extraction. Water solvent debinding was carried out at three temperatures stepwise during 5 h. As a result, efficient removal of the PEG as well as free defects samples were obtained after solvent debinding for binder systems based on low molecular weight of PEG.     

Poly(ethylene oxide) Grafted to Silicon Surfaces: Grafting Density and Protein Adsorption

Poly(ethylene oxide) (PEO) polymer, in linear and star form, was covalently grafted to silicon surfaces, and the surfaces were tested for their ability to adsorb proteins. Linear PEG of molecular weight 3400, 10 000, and 20 000 g/mol and star PEO molecules were coupled via their terminal hydroxyl groups activated by tresyl chloride to aminosilane-treated silicon wafers. The amount of PEO coupled to the surface was varied by changing the concentration of the tresyl-PEO solution. The dry PEO thickness on the surface was measured using X-ray photoelectron spectroscopy (XPS) and ellipsometry, from which the grafting density was calculated. The PEO surfaces were exposed to solutions of each of three proteins: cytochrome-c, albumin, and fibronectin. The degree of adsorption of each protein was determined by XPS and ellipsometry and recorded as a function of PEO grafting density. All three proteins were found to reach zero adsorption at the highest grafting densities on all three PEG surfaces, which for all three PEG surfaces was a PEO content of 100 +/- 10 ng/cm2. On both star PEO surfaces, albumin and fibronectin decreased to zero adsorption at intermediate values of grafting density, whereas cytochrome-c continued to adsorb at all grafting densities, although with a decreasing trend. A physical model of the surface helped explain these protein adsorption results in terms of the spacing and degree of overlap of grafted PEO chains.     

Hydrogels of poly(ethylene glycol): mechanical characterization and release of a model drug

Thermosensitive polymer networks were synthesized from poly(ethylene glycol), hexamethylene diisocyanate and 1,2,6-hexanetriol in stoichiometric proportions. By varying the amount of 1,2,6-hexanetriol and the molar mass of the poly(ethylene glycol), a wide range of networks with different crosslinking densities was prepared. The networks obtained were characterized by the temperature dependence of their degree of equilibrium swelling in water and by their Young's moduli. For each network, the molecular weight between crosslinks was estimated. The structure of the hydrogels was analysed with respect to scaling laws, and it was found that the results obtained with PEG 1500 and PEG 6000 hydrogels are in agreement with theoretical predictions, whereas those obtained with PEG 400 hydrogels are in disagreement. The release properties of PEG hydrogels were studied by the determination of the diffusion coefficient for acebutolol chlorhydrate and by an analysis of the effect of temperature on these coefficients. Finally, these release properties were correlated with the swelling and structural properties of the hydrogels.     

Photodynamic effect of polyethylene glycol-modified fullerene on tumor

Fullerence (C60) efficiently generates singlet oxygen when irradiated with light, and thus should have a photodynamic effect on tumors, if it is accumulated in the tumor tissue. To explore tumor targeting of C60, we chemically modified the water-insoluble C60 with polyethylene glycol (PEG), not only to make it soluble in water, but also to enlarge its molecular size. When injected intravenously into mice carrying a tumor mass in the back subcutis, the C60-PEG conjugate exhibited higher accumulation and more prolonged retention in the tumor tissue than in normal tissue. The conjugate was excreted without being accumulated in any specific organ. Following intravenous injection of C60-PEG conjugate or Photofrin to tumor-bearing mice, coupled with exposure of the tumor site to visible light, the volume increase of the tumor mass was suppressed and the C60 conjugate exhibited a stronger suppressive effect than Photofrin. Histological examination revealed that conjugate injection plus light irradiation strongly induced tumor necrosis without any damage to the overlying normal skin. The antitumor effect of the conjugate increased with increasing irradiation power and C60 dose, and cures were achieved by treatment with a dose of 424 micrograms/kg at an irradiation power of 107 J/cm2. These findings indicate that PEG-modified C60 is a candidate agent for photodynamic tumor therapy.     

Biomolecular modification of p(AAm-co-EG/AA) IPNs supports osteoblast adhesion and phenotypic expression

Interpenetrating polymer networks (IPNs) were designed to resist materials fouling caused by non-specific protein adsorption, and indiscriminate cell or bacterial adhesion. These IPNs were thin adherent films (approximately 20 nm) comprised of acrylamide (AAm), ethylene glycol (EG), and acrylic acid (AA) grafted to either silicon waters or quartz substrates via photoinitiated free radical polymerization. These networks were further modified to promote specific cell adhesion by tethering bioactive groups such as peptides that mimic cell-binding domains found on extracellular matrix molecules. As a specific example of biomolecular surface engineering, peptides from the cell-binding domain of bone sialoprotein were tethered to a p(AAm-co-EG/AA) IPN to control cell behavior at the surface. The networks were characterized by contact angle measurements, spectroscopic ellipsometry, and X-ray photoelectron spectroscopy to convey information on IPN wettability, thickness, and chemistry. The surface characterization data supported the theory that the PEG/AA layer formed an IPN with the underlying p(AAm) network, and after graft modification of this IPN with diamino PEG (PEG(NH2)2), the PEG(NH2)2 chains were enriched at the surface. Rat calvarial osteoblasts attached to Arg-Gly-Asp (RGD) modified IPNs at levels significantly greater than on clean quartz, Arg-Gly-Glu (RGE) modified, or the PEG(NH2)2 modified IPN, with or without serum in the media. Cells maintained in media containing 15% fetal bovine serum (FBS) proliferated, exhibited nodule formation, and generated sheets of mineralized extracellular matrix (ECM) with the addition on beta-glycerophosphate to the media. Cell adhesion and mineralized ECM formation were specifically dependent on the peptide sequence present at the surface.     

Characterization of Ceramic Composite-Membranes Prepared by ORMOSIL Coating Sol

Sol-gel methods offer many advantages over conventional slip-casting, including the ability to produce ceramic membranes. They are purer, more homogeneous, more reactive and contain a wider variety of compositions. We produced ormosil sol using sol-gel process under different molecular weight of polymer species [polyethylene glycol (PEG) ] in total system [Tetraethyl ortho silicate(TEOS)-polyethylene glycol (PEG)]. The properties of as-prepared ormosil sol such as,viscosity, gelation time were characterized. Also, the ceramic membrane was prepared by dip-coating with synthetic sol and its micro-structure was observed by scanning electron microscopy. The permeability and rejection efficiency of membrane for oil/water emulsion were evaluated as cross-flow apparatus. The ormosil sol coated Membrane is easily formed by steric effect of polymer and it improves flux efficiency because infiltration into porous support decreased. Its flux efficiency is elevated about 200(1/m2·h) compared with colloidal sol coated membrane at point of five minutes from starting test.     

Erosion of biodegradable block copolymers made of poly(D,L-lactic acid) and poly(ethylene glycol)

Biodegradable block copolymers made of poly(ethylene glycol) monomethylether (Me.PEG) and poly(D,L-lactic acid) (PLA) were investigated for their erosion properties. Wide angle X-ray diffraction (WAXD) and differential scanning calorimetry (DSC) investigations prior to erosion revealed that despite the low content of crystallizable Me.PEG of 10%, Me.PEG5-PLA45 is a partially crystalline polymer. The erosion of the polymer was investigated using cylindrical polymer matrix discs with a diameter of 8 mm and a height of 1.5 mm. WAXD and DSC spectra obtained from eroded polymer matrix discs suggest that both polymer blocks separate completely during erosion. The crystallinity of Me.PEG5-PLA45 was found to increase during erosion, which is probably due to the improved mobility of Me.PEG inside the polymer with a progressive degree of degradation. The erosion kinetics were found to be similar to that of PLA or poly(lactic-co-glycolic acid). During erosion the polymer matrix weight of dried samples remains constant for 11 weeks after which erosion sets in rapidly. From this observation one can conclude that the impact of the relatively small Me.PEG chains on Me.PEG5-PLA45 erosion is not pronounced. This is beneficial for all those applications that require the stability of the polymer matrix and in which the Me.PEG chain is intended to bring about other effects such as the modification of the surface properties of PLA polymers.     

Endgroup analysis of polyethylene glycol polymers by matrix-assisted laser desorption/ionization Fourier-transform ion cyclotron resonance mass spectrometry

Fourier-transform ion cyclotron resonance mass spectrometry (FTICR-MS) by external injection of matrix-assisted laser desorbed and ionized (MALDI) polymers offers good possibilities for characterization of low molecular weight homopolymers (MW range up to 10 kDa). The molecular masses of the molecular weight distribution (MWD) components of underivatized and derivatized (dimethyl, dipropyl, dibutyl and diacetyl) polyethylene glycol (PEG) 1000 and 4000 were measured by MALDI-FTICR-MS. These measurements have been performed using a commercial FTICR spectrometer with a home-built external ion source. MALDI of the samples with a 2,5-dihydroxybenzoic acid matrix in a 1000:1 matrix-to-analyte molar ratio produces sodiated molecules in a sufficient yield to trap the ions in the ICR cell. The masses of the molecular weight distribution of PEG components were measured in broad-band mode with a mass accuracy of < 5 ppm in the mass range around 1000 u and within 40 ppm accuracy around 4000 u. From these measurements, the endgroup mass of the polymer was determined by correlation of the measured component mass with the degree of polymerization. The masses of the PEG endgroups have been determined within a deviation of 3-10 millimass units for the PEG1000 derivatives and 10-100 millimass units for the PEG4000 derivatives, thus confirming the identity of the distal parts of the model compounds.     

Preparation and characterization of poly(propylene fumarate-co-ethylene glycol) hydrogels

We describe the preparation and bulk characterization of a cross-linked poly(propylene fumarate-co-ethylene glycol), p(PF-co-EG), hydrogel. Eight block copolymer formulations were made varying four different design parameters including: poly(ethylene glycol) (PEG) molecular weight, poly(propylene fumarate) (PPF) molecular weight, copolymer molecular weight, and ratio of PEG to PPF. Two different cross-linking formulations were also tested, one with a cross-linking monomer and one without. The extent of the cross-linking reaction and the degree of swelling in aqueous solution were determined on copolymer formulations made without a cross-linking monomer. The values of molecular weight between cross-links, Mc ranged from 300 +/- 120 to 1190 +/- 320 as determined from swelling data (n = 3). The equilibrium volume swelling ratios, Q, varied from 1.5 +/- 0.1 to 3.0 +/- 0.1. This ratio was found to increase with increasing PEG content in the copolymer and decrease with increasing PPF molecular weight. The values for complex dynamic elastic moduli magnitudes of E*, ranged from 0.9 +/- 0.2 to 13.1 +/- 1.1 MPa for the formulations with the cross-linking monomer, N-vinyl pyrrolidinone (VP) (n = 3). The ultimate tensile stresses on the formulations made with VP ranged from 0.15 +/- 0.03 to 1.44 +/- 1.06 MPa, and tensile moduli ranged from 1.11 +/- 0.20 to 20.66 +/- 2.42 MPa (n = 5). All of the mechanical properties increased with increasing PPF molecular weight and decreased with increasing PEG content in the copolymer. These data show that the physical properties of p(PF-co-EG) hydrogels can be tailored for specific applications by altering the material composition.     

Endoscopic brain surgery of intracranial cysts: report of 2 cases

It is well established that polyethylene glycol (PEG) shifts the equilibrium in oligomeric protein systems to form higher molecular weight associates. We used this effect of PEG to evaluate a modification of functional properties of LDH from pig skeletal muscles. PEG decreases the rate of heating-induced LDH inactivation in the concentration dependent manner. Michaelis constant and maximal velocity of the enzyme as well as inhibition of LDH by high pyruvate concentrations were affected by PEG. Enzyme preincubation with PEG suppresses also the formation of ternary inactive complex NAD-pyruvate-LDH.     

Interaction of poly(ethylene-glycols) with air-water interfaces and lipid monolayers: investigations on surface pressure and surface potential

We have characterized the surface activity of different-sized poly(ethylene-glycols) (PEG; M(r) 200-100,000 Da) in the presence or absence of lipid monolayers and over a wide range of bulk PEG concentrations (10(-8)-10% w/v). Measurements of the surface potential and surface pressure demonstrate that PEGs interact with the air-water and lipid-water interfaces. Without lipid, PEG added either to the subphase or to the air-water interface forms relatively stable monolayers. Except for very low molecular weight polymers (PEGs < 1000 Da), low concentrations of PEG in the subphase (between 10(-5) and 10(-4)% w/v) increase the surface potential from zero (with respect to the potential of a pure air-water interface) to a plateau value of approximately 440 mV. At much higher polymer concentrations, > 10(-1)% (w/v), depending on the molecular weight of the PEG and corresponding to the concentration at which the polymers in solution are likely to overlap, the surface potential decreases. High concentrations of PEG in the subphase cause a similar decrease in the surface potential of densely packed lipid monolayers spread from either diphytanoyl phosphatidylcholine (DPhPC), dipalmitoyl phosphatidylcholine (DPPC), or dioleoyl phosphatidylserine (DOPS). Adding PEG as a monolayer at the air-water interface also affects the surface activity of DPhPC or DPPC monolayers. At low lipid concentration, the surface pressure and potential are determined by the polymer. For intermediate lipid concentrations, the surface pressure-area and surface potential-area isotherms show that the effects due to lipid and PEG are not always additive and that the polymer's effect is distinct for the two lipids. When PEG-lipid-mixed monolayers are compressed to surface pressures greater than the collapse pressure for a PEG monolayer, the surface pressure-area and surface potential-area isotherms approach that of the lipid alone, suggesting that for this experimental condition PEG is expelled from the interface.     

Spatially controlled cell engineering on biodegradable polymer surfaces

Controlling receptor-mediated interactions between cells and template surfaces is a central principle in many tissue engineering procedures (1-3). Biomaterial surfaces engineered to present cell adhesion ligands undergo integrin-mediated molecular interactions with cells (1, 4, 5), stimulating cell spreading, and differentiation (6-8). This provides a mechanism for mimicking natural cell-to-matrix interactions. Further sophistication in the control of cell interactions can be achieved by fabricating surfaces on which the spatial distribution of ligands is restricted to micron-scale pattern features (9-14). Patterning technology promises to facilitate spatially controlled tissue engineering with applications in the regeneration of highly organized tissues. These new applications require the formation of ligand patterns on biocompatible and biodegradable templates, which control tissue regeneration processes, before removal by metabolism. We have developed a method of generating micron-scale patterns of any biotinylated ligand on the surface of a biodegradable block copolymer, polylactide-poly(ethylene glycol). The technique achieves control of biomolecule deposition with nanometer precision. Spatial control over cell development has been observed when using these templates to culture bovine aortic endothelial cells and PC12 nerve cells. Furthermore, neurite extension on the biodegradable polymer surface is directed by pattern features composed of peptides containing the IKVAV sequence (15, 16), suggesting that directional control over nerve regeneration on biodegradable biomaterials can be achieved.     

Light-induced tailoring of PEG-hydrogel properties

We have previously reported (Andreopoulos et al. J Am Chem Soc 118 (1996) 6235-6240) the synthesis of hydrogels via the photopolymerization of water-soluble PEG molecules. In this paper, PEG-hydrogel membranes were prepared by the irradiation (> 300 nm) of aqueous solutions of photosensitive 4-armed PEG (nominal molecular weight of 20000), in the absence of photo-initiators. The hydroxyl termini of the PEG's were functionalized with cinnamylidene acetate groups to form photosensitive PEG macromers (PEG-CA), which upon irradiation (>300 nm) formed crosslinks between adjacent cinnamylidene groups resulting in highly crosslinked networks (hydrogels) (Andreopoulos et al. J Am Chem Soc 118 (1996) 6235-6240). The hydrogel membranes were highly swellable with equilibrium volume fractions ranging from 0.02 to 0.05. Their swellability was a function of irradiation light (>300 nm) and degree of modification of the PEG molecules. The effect of light on the permeation fluxes of myoglobin (Mb), hemoglobin (Hb), and lactate dehydrogenase-L (LDH) through PEG membranes was also assessed and the diffusion coefficients of the proteins were determined accordingly. The PEG-CA membranes exhibited photoscissive behavior upon exposure to UV irradiation (254 nm). Therefore, UV light was used as a trigger to control the mesh size of the membranes, and thereby the permeation fluxes of Mb, Hb, and LDH. Equilibrium swelling experiments with membranes prepared under different irradiation conditions were performed, and the Flory-Huggins model was utilized to determine the mesh size and the average molecular weight between crosslinks of the synthesized hydrogels.     

Characterization of permeability and network structure of interfacially photopolymerized poly(ethylene glycol) diacrylate hydrogels

Hydrogel membranes formed by interfacially photopolymerizing poly(ethylene glycol) (PEG) diacrylate precursor solution were prepared from PEG diacrylate of molecular weights (MW) ranging from 2000 (2K) to 20000 (20K) with concentrations ranging from 10% to 30% w/w. The effects of PEG diacrylate MW and concentration in the membrane precursor solution upon the diffusivities of vitamin B12, myoglobin, ovalbumin, albumin, and IgG were determined. Regardless of the concentration of the PEG diacrylate in the precursor solution, hydrogels prepared with PEG 2K, 4K, and 8K diacrylate were impermeable to proteins with a size equal to or larger than myoglobin (22 kDa), while hydrogels prepared with PEG 20K diacrylate were impermeable to proteins with a size equal to or larger than ovalbumin (45 kDa). Similarities between hydrogels formed from PEG 2K, 4K, and 8K diacrylates were also seen in calculations of the molecular weight between crosslinks and the mesh size, with values in the range of 150-750 g/mol and 15-35 A, respectively, depending on PEG diacrylate concentration. In contrast, hydrogels formed from PEG 20K diacrylate had molecular weight between crosslinks ranging from 1150 to 2000 g/mol and mesh sizes ranging from 45-70 A, with larger values being observed in membranes polymerized from more dilute PEG diacrylate precursor.     

一种新型水溶性金属注射成形用粘结剂的研制

论文在抽提MI M粘结剂中水溶性组分及高分子组元功能的基础上,通过参数计算、溶解试验确定了水溶性组分的品种、PEG的分子量分布,进而结合混合试验以及扫描电镜测试确定了水溶性MI M粘结剂中的高分子组元和整个粘结剂配方。结果表明,PEG是一种合适的水溶性组分,恰当的分子量分布,即10%PEG600+30%PEG1000+60%PEG1500的混合配方可以满足MI M工艺对其热收缩率的要求;PEG-POM具有热力学相容性,其混合物流动性、冷却强度好但水溶性差;PMMA分子量对PMMA-PEG的混合性具有决定性的作用,在PMMA分子量适中(M=(40·7~98·4)×104)时,PEG-PMMA混合物相容性、冷却强度和水溶性都较好,但流动性稍差;当PEG-POM-PMMA三者以75∶23∶2配比组成三元系时,它兼备了PEG-POM和PEG-PMMA二元体系的优点,总体效果最好。为了增加粘结剂水溶性组分PEG的热稳定性、改善组分间的相容性、降低黏度和增加粘结剂对粉末的润湿性,粘结剂需要配以适量抗氧剂BHT、增塑剂DOP和表面活性剂。最终确定粘结剂适宜配方为:PEG72%,POM23%,PMMA2%,SA2%,DOP1%,BHT0·01%。     

Role of water in protein kinase C catalysis and its binding to membranes

The role of hydration in the catalytic activity and membrane binding of rat brain protein kinase C (PKC) was investigated by modulating the activity of water with polyethylene glycols with molecular weights of 1000-20000 and dextran with a molecular weight of 20000. These polymers create an osmotic stress due to their exclusion from hydration shells and crevices on proteins, causing dehydration. Polymers larger than 1000 caused an activation of the PKC-catalyzed phosphorylation of histone, while PEG 1000 had no significant effect. The extent of activation by PEG and dextran 20000 was larger than that of PEG 6000 or 8000 when vesicles were composed of 1:1 POPS/POPC, suggesting the presence of at least two distinct regions of exclusion on PKC: one inaccessible to PEGs larger than 1000 and the other inaccessible only to PEGs of > 10000. The extent of activation was dependent on the composition of the vesicles used. If basal activity (without PEG) was low (e.g. with low PS content in membranes), then the extent of activation was similar for all polymers larger than 1000. Binding of PKC to membranes containing 50 mol % PS was unaffected by PEG 6000 but was inhibited by PEG 20000. At a low PS content of 10%, both PEG 6000 and 20000 inhibited binding. This suggests that PKC becomes hydrated upon binding to membranes. Under conditions in which all of the enzyme is membrane-bound, both Km and Vmax for the phosphorylation of histone increased linearly with osmotic stress induced by PEG 6000. Thus, PKC becomes hydrated with 2311 +/- 476 water molecules upon binding of histone and is dehydrated by 1349 +/- 882 water molecules in going to the transition state. Km and Vmax for phosphorylation of the MARCKS peptide also increase with osmotic stress induced by PEG 6000. When protamine sulfate was used as a substrate (cofactor-independent), Vmax for the reaction was unaffected, but Km decreased with osmotic pressure (with PEG 6000), suggesting that PKC becomes dehydrated upon binding protamine. Similar results were found with a peptide substrate derived from the pseudosubstrate site of PKC epsilon. Since dextran, a polymer unrelated in structure to PEG, could cause a similar activation of PKC, the effects seen are likely due to osmotic stress and not to specific binding of PEG to PKC. Also, results obtained with PE-linked PEG were opposite to those with free PEG. PE-linked PEGs of 2000 and 5000 caused an inhibition of PKC-catalyzed phosphorylation of histone when present in membranes. If a specific interaction occurred with PEG, this would be expected to occur even with PE-PEG. The effects observed with free PEG are also independent of ionic strength. Free PEG had no effect on the bilayer to hexagonal phase transition temperature of DEPE membranes, suggesting that the effects on PKC activity are not a consequence of changes in membrane properties at the osmotic pressures used.     

Colloid determination of fibrin network permeability

The effects of polymers, dextran and polyvinylpyrolidone (PVP) and of albumin on the permeability of thrombin-induced fibrin networks developed in plasma were examined. Both PVP and dextran increased the network permeability and turbidity and increased the fibrin fibre thickness. The effect was molecular weight dependent. Derivation of the dimensionless permeability (permeability/fibre radius2) indicated that the increase in network permeability was mainly from altered arrangement of fibres and not from increased fibre thickness. The effects of albumin on network structure were similar to those of the polymers. Scanning electron microscopy of networks developed in plasma under the influence of dextran and poloxamer 188 showed fibres with increased thickness and a coarse nodular appearance. There was an increased tendency for fibres to be aggregated into clumps. It is suggested that during polymerization fibrin fibres and fibrin polymerization intermediaries behave as colloidal particles. Attractive forces between the particles are generated by soluble macromolecules such as plasma proteins or polymers. Attractive forces increase the thickness of fibrin fibres and induce a more permeable arrangement of the fibres in the network. The most likely colloidal mechanism is depletion flocculation. This would account for (1) the molecular weight dependence and concentration dependence of the effects of macromolecules, (2) the effects of macromolecules which do not bind to fibrin, (3) the effects of the surfactant poloxamer 188. Depletion flocculation may be a significant mechanism for biological regulation of fibrin network permeability by non-specific macromolecules such as soluble proteins or fibrin intermediaries.     

NGF depletion reduces ipsilateral and contralateral trigeminal satellite cell reactions after inferior alveolar nerve injury in adult rats

The degradation of poly(propylene fumarate-co-ethylene glycol) hydrogels was examined in vitro in phosphate-buffered saline at pH 7.4 and in vivo in a subcutaneous rat model. These hydrogels have potential application as biodegradable, injectable cardiovascular stents, and, as such, their mass loss, dimensional changes, mechanical properties, morphology, and biocompatibility over a 12-week time course were evaluated. Three formulations were fabricated: one base formulation consisting of 25% (w/w) PEG, molecular weight 4,600; one high weight percent PEG formulation with 50% (w/w) PEG; and one high molecular weight PEG formulation, molecular weight 10,500. All three formulations showed significant weight loss (between 40 and 60%) on the first day due to leaching of the uncrosslinked fraction. Further weight loss was observed only for the low weight percent PEG copolymers in the in vivo case, and a slight increase in volume was observed due to degradative swelling. The mechanical properties of the P(PF-co-EG) hydrogels decreased significantly in the first 3 weeks, showing the biphasic pattern typical of bulk degradation. In vitro, the hydrogels showed at least a 20% retention of their initial ultimate tensile stress after 3 weeks. The dynamic mechanical properties showed similar retention, with the in vivo mechanical properties differing from the in vitro properties only after 6 weeks of degradation. Differences in PEG molecular weight appeared to have little effect, but increasing the weight percent PEG decreased the rate of degradation both in vitro and in vivo. The morphology of the copolymer films, based on scanning electron microscopy observation, was not significantly different either among the three formulations or over the time course of the study, suggesting there were no macroscopic structural changes during this time period. The P(PF-co-EG) hydrogels demonstrated good initial biocompatibility, showing responses characteristic of biomaterial implants.