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Fluoroalkylethyl silane coating as a moisture barrier for dental ceramics.

The performance of dental ceramics is reduced in wet environments due to stress-corrosion limiting restoration lifetime. As analysis suggests that fracture is typically initiated from the internal surface, placement of a thin coating before a restoration is cemented to place may prevent fracture if it acts as a moisture barrier. Of coatings we tested, a fluoroalkylethyl silane (du Pont MPD-7680) showed promise. This study aimed to test a range of concentrations on glass and to determine any strength improvement on dental ceramics. Glass slides (n = 8-9) were indented (29-N load) stored 24 h, etched, coated with 100, 50, and 25% fluorosilane in isopropyl alcohol, and air-cured for 24 h. The slides were fractured on a biaxial fixture in water at 0.5 mm/min and maximum stresses calculated. Uncoated controls were tested in water and liquid N2 to determine the inert strength without stress-corrosion. Results showed increasing mean strength from uncoated (53.6 MPa) to 100% (68.0 MPa) which compared to the 80.4 MPa inert strength. ANOVA showed P < .05. Subsequently the experiment was repeated on discs of three dental ceramics (n = 7-10): feldspathic (Ceramco), glass-ceramic (Dicor MGC), high-leucite (Optec) with the 100% solution only. Mean strength was significantly (P < .01) enhanced in the feldspathic (56.2 MPa, untreated, 70.7 MPa, treated) but not the high-leucite (83.1, 83.0) or the glass-ceramic (283.9, 271.4). Fluoroalkylethyl silane coating reduced stress-corrosion in glass and feldspathic porcelain. It may be a practical approach to reducing fracture of ceramic restorations.

Ceramics↗

Magnitude and polarity of a fluoroethylene propylene electret substrate charge influences neurite outgrowth in vitro.

Positively charged coating materials such as polylysine improve neuronal attachment in vitro. Due to the structural complexity of these charged molecules, it is unclear whether neuronal effects are due to charge or to physicochemical effects, or both. Polymeric materials with charge storage capabilities and defined surface properties may provide a model in which electrical charge and surface property effects can be separated. Fluorinated ethylenepropylene (FEP) films can store negative or positive charges injected through a corona charging process, thus generating a negative or positive external electrostatic field. In the present study, mouse neuroblastoma (Nb2a) cells were cultured on positive, negative, and uncharged FEP substrates, in both serum-containing and serum-free media. Cell attachment, differentiation, and neurite outgrowth were assessed 24, 48, 72 and 96 h after plating. Electron spectroscopy for chemical analysis (ESCA), contact angle analysis, and scanning electron microscopy (SEM) revealed no differences in surface chemistry and topography between positive, negative, and uncharged FEP. No significant differences in the levels of cell attachment on positive, negative, and uncharged substrates were observed. Significantly higher levels of neurite outgrowth, however, were observed with positive substrates as compared to negative and uncharged substrates, in both media conditions. Substrates charged to +1000 V showed greater levels of outgrowth compared to +500 and +3000 V, suggesting the presence of an optimal range of charge for neurite outgrowth. These results show that cell/charge interactions mediate cell effects on electrically charged substrates with identical surface chemistry, topography and adhesivity.

Animals↗

Synthesis and characterization of fluoropolymeric substrata with immobilized minimal peptide sequences for cell adhesion studies. I.

In this work, poly(tetrafluoroethylene-co-hexafluoropropylene) (also known as fluorinated ethylene propylene; FEP) was functionalized at the surface using a radio frequency glow discharge plasma. This particular surface modification produced controlled densities of hydroxyl functionality on the FEP surface. These surface hydroxyl groups provided sites for the covalent attachment of minimal peptide sequences, that are specific for neuronal attachment. FSCA, ATR-FTIR, ToF-SIMS, and fluorescence spectroscopy were used to evaluate peptide reaction efficiencies and to verify that intact peptide sequences were covalently attached to the FEP surfaces. These modified substrata were then used to study the cell attachment and response to covalently bound minimal peptide sequences. Cell attachment and differentiation results using NG108-15 and PC12 neuronal cell lines are presented in the adjoining paper by Ranieri et al.

Amino Acid Sequence↗

Neuronal cell attachment to fluorinated ethylene propylene films with covalently immobilized laminin oligopeptides YIGSR and IKVAV. II.

Material surfaces that can mediate cellular interactions by the coupling of specific cell membrane receptors may allow for the design of a biomaterial that can control cell attachment, differentiation, and tissue organization. Cell adhesion proteins have been shown to contain minimum oligopeptide sequences that are recognized by cell surface receptors and can be covalently immobilized on material surfaces. In this study, cell attachment to fluorinated ethylene propylene (FEP) films functionalized with the laminin-derived oligopeptides, YIGSR and a 19-mer IKVAV-containing sequence, was assessed using NG108-15 neuroblastoma and PC12 cells. A radiofrequency glow discharge (RFGD) process that replaces the FEP surface fluorine atoms with reactive hydroxyl functionalities was used to activate the film surfaces. The oligopeptides were then covalently coupled to the surface by their C-terminus using a standard nucleophilic substitution reaction. The covalent attachment of the oligopeptides to the FEP surface was verified using electron spectroscopy for chemical analysis (ESCA). Receptor-mediated NG108-15 cell attachment on the YIGSR-modified films was determined using competitive binding assays. Average cell attachment on the oligopeptide immobilized films in medium containing soluble CDPGYIGSR was reduced by approximately a factor of 2, compared to cell attachment in serum-free medium alone. No significant decrease in cell attachment was noted in medium containing the mock oligopeptide sequence CDPGYIGSK. FEP films immobilized with the 19-mer IKVAV sequence demonstrated a higher percentage of receptor mediated cell attachment on the film surfaces.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

Polyvinylidene fluoride (PVDF) as a biomaterial: from polymeric raw material to monofilament vascular suture.

This study identified the effects of various manufacturing processes on the crystalline microstructure, mechanical properties, and biocompatibility of a polyvinylidene fluoride (PVDF) suture. To achieve this, changes in the crystalline microstructure and the tensile behavior of PVDF monofilaments were monitored in vitro after different thermal processing, coloration, and sterilization treatments. In addition, the in vivo biocompatibility of the manufactured and sterilized PVDF suture was assessed by using it to anastomose a preclotted polyester vascular prosthesis as a thoracoabdominal bypass in a series of dogs. The tissue response was followed by histologic and scanning electron microscopy over implantation periods ranging from 4 h to 6 months. Differential scanning calorimetry and infrared spectroscopy (FTIR-ATR) showed that thermal processing and the addition of a coloring agent had a direct effect on modifying the crystalline microstructure and hence changing the mechanical properties. For example, thermal processing converted some of the alpha phase into the beta and gamma polymorphs, whereas coloration led only to a major increase in the beta-to-alpha ratio. The tensile properties were found to be optimized when the relative proportion of the beta and gamma phases combined compared to the alpha form gave rise to an FTIR A509/A532 absorption ratio between 4.0 and 4.5. Sterilization was found to cause some modifications to the crystalline microstructure near the surface of the monofilaments, but it did not change their mechanical properties. Pathologic examination of the anastomotic regions after different periods of implantation revealed a minimal cellular response, with no mineralization, intimal hyperplasia, or excessive fibrous tissue reaction. This good biocompatibility, together with other desirable characteristics such as ease of manipulation and satisfactory mechanical strength, makes PVDF an attractive alternative monofilament suture material for cardiovascular surgery.

Animals↗

Interactions of acinar cells on biomaterials with various surface properties.

The purpose of this study is to evaluate the interactions of rat parotid acinar cells on biomaterials with different surface properties. The biomaterials used in this study included polyvinyl alcohol (PVA), chitosan, poly (ethylene-co-vinyl alcohol) (EVAL), and polyvinylidene fluoride (PVDF). Cell morphology was observed by photomicroscope. Cell growth and differentiated characteristic function were separately assayed with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) reduction activity and amylase activity. Results indicated that behaviors of acinar cells on materials might differ to a great extent depending on the surface hydrophilicity and morphology of the materials. On the relatively hydrophobic materials, the abilities of acinar cells to adhere and proliferate increased simultaneously. In addition, porous PVDF had higher cell growth compared with dense PVDF. Therefore, the hydrophobic PVDF with a porous structure was the best substrate for culturing acinar cells. According to our findings, a tubular PVDF scaffold with dense outer surface to prevent saliva leakage and with porous inner surface for the cell growth was proposed to serve as an artificial salivary gland for future use in the treatment of patients with salivary hypofunction.

Amylases↗

In vivo biostability of polyether polyurethanes with fluoropolymer and polyethylene oxide surface modifying endgroups; resistance to metal ion oxidation.

Polyether polyurethanes are subject to oxidation catalyzed by, and through direct (redox) reaction with transition metal ions (metal ion oxidation, MIO). The source of the ions is corrosion of metallic parts within an implanted device. A Shore 80A polyether polyurethane was modified with fluoropolymer (E80AF) or polyethylene oxide (E80AP) surface modifying end groups (SME). The SME migrates to the surface to form a covalently bonded monolayer, while maintaining the bulk properties of the polyurethane. In vitro tests in H(2)O(2) solution indicated that both SME's accelerated MIO. Tubing samples containing cobalt mandrels were implanted in the subcutis of rabbits for up to 2 years. In vivo, E80AF significantly slowed the rate of visible degradation, but did not prevent MIO. E80AP had virtually identical visual performance to the unmodified control in vivo. Infrared spectroscopy and molecular weight correlated well with visual appearance. When cracks were seen, polyether soft segment oxidation was occurring. Both E80AP and the control developed severe loss of molecular weight in vivo. The changes were much less severe for E80AF. Thus, contrary to in vitro test results, the PEO SME had no effect at all on MIO resistance, while the fluoropolymer SME produced a significant improvement in biostability.

Animals↗

Fibrinogen adsorption and platelet lysis characterization of fluorinated surface-modified polyetherurethanes.

A polyetherurethane (PU) was modified using fluorinated surface-modifying macromolecules (SMMs). A double radiolabel method was used simultaneously to measure the number of adhered platelets ((51)Cr) and the quantity of adsorbed Fg ((125)I), in a cone-and-plate instrument. The objectives were to determine if adsorbed Fg levels correlated to platelet adhesion on the surfaces, and to assess if any reductions in platelet adhesion for the SMM-treated surfaces resulted from surface-induced platelet lysis, rather than changes directly related to lower platelet activation and attachment on the novel surfaces. Platelet lysis was determined from lactate dehydrogenase (LDH) and unbound (51)Cr released into plasma isolated from whole blood exposed to test materials. The corresponding Fg adsorption, evaluated under the same platelet adhesion conditions, did not account for the reduced platelet adhesion on the treated surfaces. LDH and (51)Cr platelet release were very low and indicated no statistically significant differences between the materials. It was therefore concluded that platelet lysis did not contribute to the reduction in platelet adhesion characteristic observed on the SMM-treated surfaces. More importantly, the work emphasizes that the platelet activation cannot be inferred to by assessing the quantity of fibrinogen as is commonly done in the literature. The finding suggests a much more complex mechanism of action for the SMM surface modifiers. On-going work is investigating other Fg parameters such as protein binding affinity and protein conformational state in order to establish the mechanism by which the fluorinated surface modifiers may be reducing platelet adhesion via intermediary changes in initial protein adsorption.

Adsorption↗

Liver alcohol dehydrogenase immobilized on polyvinylidene difluoride.

A physical method for immobilization of liver alcohol dehydrogenase (ADH) by hydrophobic adsorption onto a supporting membrane of polyvinylidene difluoride (PVDF) was performed. Simultaneously, a physicochemical characterization of the immobilized enzyme regarding its kinetic behaviour was performed. The activity/pH profile observed points to an effect of pH on activity that is completely different from the case of ADH in solution. The disturbance in the typical bell-shaped profile owing to the fact that the enzyme was immobilized is explained on the basis of a potent limitation to the diffusion of the protons in the support. The findings of the present work also reveal the existence of an effect that limits free external diffusion of the substrate towards and/or the product from the support; this effect seems to be the determinant of the overall rate of the enzymatic reaction and is thus of great importance in the effective kinetic behaviour (v([S])) of immobilized ADH, whose kinetic behaviour is complex (non-Michaelian), as may be seen from the lack of linearity observed in the corresponding double reciprocal and Eadie-Hofstee plots. By non-linear regression numerical analysis of the v([S]) data and application of the F-test for model discrimination, the minimum rate equation necessary to describe the intrinsic kinetic behaviour of PVDF-immobilized ADH proved to be one of the polynomial quotient type of degree 2:2 (in substrate concentration).

Alcohol Dehydrogenase↗

Piezoelectric internal fixation devices: a new approach to electrical augmentation of osteogenesis.

Prototype testing has been accomplished on a piezoelectric, internal fixation plate. This device combines a piezoelectric material with an internal fixation device as an integrated structure that provides mechanical stability, together with self-generated electrical stimulation, for treating fractures and nonunion. In bench and animal tests we have demonstrated that cyclical loading can cause a device of this type to generate electrical charge while attached to bone. After rectification, direct currents within the range known to stimulate osteogenesis can be produced by weight-bearing loads. Furthermore, electrical output of the implants can be increased by externally applied ultrasonic energy. These twin developments add significantly to the potential armamentarium of devices to enhance bone healing.

Animals↗

Occurrence, extent, and implications of pressure waves during excimer laser ablation of normal arterial wall and atherosclerotic plaque.

Ablation of atherosclerotic plaque and normal arterial wall was performed using a Xenon-Chloride Excimer laser with a wave-length of 308 nm and a pulse duration of 115 ns. The light was transmitted via a 600 microns bare fibre and adjusted to an energy density of 3.5J/cm2. The acoustic signals generated by the laser pulse were measured with two types of hydrophones consisting of polyvinylidenefluoride with active diameters of 0.3 mm and 0.5 mm and recorded on a dual channel digital storage oscilloscope using either a 0.5 m coaxial cable or a broadband fibre-optic transmission system. Tissue was retrieved from nine cadaver human aortas and macroscopically classified as either normal or calcified atherosclerotic plaque. Histological analysis (Haematoxylin eosin, elastica van Gieson, and immunohistochemical staining) was carried out after the experiments to verify the macroscopic diagnosis and to correlate the acoustic responses with the tissue characteristics. For normal arterial wall, maximum peak pressure was 1.28 MPa +/- 0.85 MPa, rise time 163 ns +/- 43 ns, and pressure increase 8.2k Pa +/- 5.4k Pa/ns. For calcified, atheromatous segments, a maximum peak pressure of 2.02 MPa +/- 1.16 MPa, a rise time of 69.9 ns +/- 25.8 ns, and a pressure increase of 32.3 kPa +/- 21.3 kPa/ns was found. Statistical analysis showed a significant shorter rise time (P < 0.0001) and a higher pressure increase (P < 0.0001) for calcified tissue in comparison to normal arterial wall, whereas maximum pressures alone did not allow a differentiation of tissue characteristics. Several hundred kPa are generated during Excimer laser ablation.(ABSTRACT TRUNCATED AT 250 WORDS)

Acoustics↗

Formation of pressure waves during in vitro excimer laser irradiation in whole blood and the effect of dilution with contrast media and saline.

Pressure waves during excimer laser ablation of vascular tissue may be responsible for complications of coronary excimer laser angioplasty. In this experimental study, pressure waves were measured during excimer laser irradiation in blood and contrast media using a polyvinilidenefluoride hydrophone. At a distance of 4 mm lateral to the tip of a 1.7 mm multifiber laser catheter, excimer laser irradiation in blood resulted in a linear increase of peak pressures from 1,365 +/- 165 kPa at 30 mJ/mm2 to 2,866 +/- 404 kPa at 60 mJ/mm2. In contrast media, peak pressure increased from 3,172 +/- 573 kPa (30 mJ/mm2) to 5,763 +/- 467 kPa (60 mJ/mm2). Contrast media and saline were added to blood. At a concentration of 60% contrast in blood, a 3.4 fold increase of peak pressures was documented as compared to pure blood. Further increase of the concentration did not result in higher pressure waves. Concentrations of saline in blood of 90% and 96% reduced the peak pressures by 16% and > 50%, respectively, as compared to pure blood.

Angioplasty, Laser↗

Proteomic analysis of acrylamide gel separated proteins immobilized on polyvinylidene difluoride membranes following proteolytic digestion in the presence of 80% acetonitrile.

Two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) combined with mass spectrometry (MS) is a highly accurate and sensitive means of identifying proteins. We have developed a novel method for digesting proteins on polyvinylidene difluoride (PVDF) membranes for subsequent matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) MS analysis. After Tricine sodium dodecyl sulfate (SDS)-PAGE, separated proteins were electroblotted onto PVDF membranes in a semidry discontinuous buffer system, visualized by staining with Coomassie Blue, excised, digested with trypsin or lysC in 80% acetonitrile, and then analyzed by MALDI-TOF MS. This method has several advantages over in-gel digestion in terms of sample handling, sensitivity, and time. We identified 105 fmol of Bacillus subtilis SecA and 100 approximately 500 fmol of standard proteins. We also analyzed the submembrane protein fraction solubilized by 1% n-dodecyl-beta-D-maltoside from B. subtilis membranes after separation by 2-D PAGE, and identified 116 protein spots. This method can detect proteins at the 10 approximately 50 fmol level by pooling more than ten identical electroblotted protein spots.

Acetonitriles↗

Profiling of the cell surface proteome.

The in depth-mining of the proteome necessitates the comprehensive analysis of proteins in individual subcellular compartments to uncover interesting patterns of protein expression that include assessment of protein location, trafficking and of post-translational modifications that are location specific. One of the compartments of substantial interest from a diagnostic and therapeutic point of view is the plasma membrane which contains intrinsic membrane proteins and other proteins expressed on the cell surface. Technologies are currently available for the comprehensive profiling of the cell surface proteome that rely on protein tagging of intact cells. Studies are emerging that point to unexpected patterns of expression of specific proteins on the cell surface, with a common occurrence of proteins previously considered to occur predominantly in other compartments, notably the endoplasmic reticulum. The profiling of the cell surface and plasma membrane proteomes will likely provide novel insights and uncover disease related alterations.

ADP-ribosyl Cyclase↗

Proteins of calcified endoskeleton: II partial amino acid sequences of endoskeletal proteins and the characterization of proteinaceous organic matrix of spicules from the alcyonarian, Synularia polydactyla.

Calcified organic substances in the skeleton contain a protein-polysaccharide complex taking a key role in the regulation of bio-calcification. However, information concerning the matrix proteins in alcyonarian and their effect on calcification process is still unknown. For this reason, we have studied the organic matrix of endoskeletal spicules from the alcyonarian coral, Synularia polydactyla, to analyze the proteins with their sequences and investigate the functional properties by a molecular approach. The separated spicules from the colony were identified by scanning electron microscope (SEM). The soluble organic matrix comprised 0.04% of spicule weight. By recording decline of pH in the experimental design, the inhibitory effect of the matrix on CaCO3 precipitation was revealed. Prior to electrophoresis, our analysis of proteins extracted from the soluble organic matrix of the spicules revealed an abundance of proteins in molecular weight. The sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis of the preparations showed seven bands of proteins with an apparent molecular mass of 109, 83, 70, 63, 41, 30 and 22 kDa. The proteins were electrophoresed on Tricine-SDS-PAGE after electro-elution treatment, and then transferred to polyvinylidene difluoride (PVDF) membranes and their N-termini were sequenced. Two bands of proteins of about 70 and 63 kDa successfully underwent N-terminal amino acid sequencing. For the detection of calcium binding proteins, a Ca2+ overlay analysis was conducted on the extract by 45Ca autoradiography. The 109 and 63 kDa calcium binding proteins were found to be radioactive. Periodic acid schiff staining indicated that 83 and 63 kDa proteins were glycosylated. An assay for carbonic anhydrase, which is thought to play an important role in the process of calcification revealed low level of the activity. These findings suggest that the endoskeletal spicules of alcyonarian corals have protein-rich organic matrices, which might be related to the calcification process.

Amino Acid Sequence↗