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Layer-by-layer assembly of polyelectrolyte films improving cytocompatibility to neural cells.

The using of layer-by-layer assembly polyelectrolyte (PE) films has been suggested as a new versatile technique for surface modification aimed at tissue engineering and cell-based chips. In this study, we investigated the surface morphology of the hyaluronic acid (HA)-based PE films deposited on the amino-functionalized glass slides using atomic force microscopy. These thin films (bilayer number <9) were measured to have nanoscale roughness ranging from 10 to 100 nm. Then the primary hippocampal and cortical neural cells were cultured on the PE films, respectively. After 5 days of culturing, the cytocompatibility to neural cells was evaluated by cellular morphology, neurite outgrowth, and microtubule-associated protein 2 expressions. From the present results, the HA-based PE films were found to be able to support neural cell adhesion and neurite development, especially for the polycation-ending films. It is suggested these HA-based multilayer PE films or similar build-ups could thus be used in the future as a way to modify surfaces for nerve scaffolds and neuron-based chips.

Animals↗

Expression of leukocyte-endothelial cell adhesion molecules on monocyte adhesion to human endothelial cells on plasma treated PET and PTFE in vitro.

We used a coculture model to evaluate the inflammatory potential of ammonia gas plasma modified PET and PTFE by flow cytometry and immunohistochemistry. In these studies, human endothelial cells from umbilical cord (HUVEC) and promonocytic U937 cells were used. HUVECs grown on polystyrene tissue culture coverslips and HUVECs stimulated with tumour necrosis factor (TNF-alpha) were used as controls. U937 adhesion to endothelium on each surface was evaluated at day 1 and day 7. To further investigate the role of leukocyte-endothelial cell adhesion molecules (CAMs) in cell-to-cell interaction on material surfaces, the expression of the leukocyte-endothelial CAMs: ICAM-1, VCAM-1, PECAM-1, and E-selectin on HUVECs were evaluated after U937 cell adhesion. The results demonstrated that plasma treated PET (T-PET) and treated PTFE (T-PTFE) did not increase U937 cell adhesion compared to the negative control. Maximal adhesion of U937 cells to HUVEC was observed on TNF-alpha stimulated endothelium with significant differences between day 1 and day 7, which is consistent with our prior observation that T-PET and T-PTFE did not cause HUVECs to increase the expression of adhesion molecules. After U937 cell adhesion, the expression of ICAM-1 and VCAM-1 of HUVECs were not different on T-PET and T-PTFE compared with the negative control. However, the expression of E-selectin was reduced on day 1, but not on day 7. The effects of plasma treated PET and PTFE on HUVEC adhesion and proliferation were also studied. On day 1 there were slight increases in the growth of HUVECs on both of T-PET and T-PTFE but this was not statistically significant. On day 7, the cell number increased significantly on the surfaces compared to the negative control. The results demonstrate that the plasma treatment of PET and PTFE with ammonia improves the adhesion and growth of endothelial cells and these surfaces do not exhibit a direct inflammatory effect in terms of monocyte adhesion and expression of leukocyte-endothelial CAMs. The monocyte adhesion to endothelial cells on surfaces can be used as a tool for the evaluation of material surface modification and further to study the mechanisms of cell-to-cell interactions in response to surfaces.

Biocompatible Materials↗

Field-effect transistor array for monitoring electrical activity from mammalian neurons in culture.

A field-effect transistor (FET) array has been fabricated and used for recording of electrical signals from neural cells. The array consists of p-channel FETs with non-metalized gates. The size of the gates of the 16 FETs are from 28 x 12 microns2 down to 10 x 4 microns2 and are arranged in a 4 x 4 matrix on 200 microns centers. For the device fabrication process we have especially focused on high sensitivity, good long-term stability in physiological conditions, and sufficient reduced signal-to-noise ratio. Special care was taken on the encapsulation technique of the device to allow surface modification based on the self-assembly technique. It can be shown that the microelectronic device surface can be modified with a synthetic peptide linked to the surface. Tailoring of the surface composition using this method allows hippocampal neurons to adhere and grow for days. More importantly, these cells develop typical electrical characteristics when cultured on this artificial surface. Using this approach neuron-FET couplings were recorded.

Animals↗

Development of a nonviral gene delivery vehicle for systemic application.

Polycation vehicles used for in vitro gene delivery require alteration for successful application in vivo. Modification of polycations by direct grafting of additional components, e.g., poly(ethylene glycol) (PEG), either before or after DNA complexation, tend to interfere with polymer/DNA binding interactions; this is a particular problem for short polycations such as linear, beta-cyclodextrin-containing polycations (betaCDPs). Here, a new method of betaCDP polyplex (polycation/DNA composite structures) modification is presented that exploits the ability to form inclusion complexes between cyclodextrins and adamantane. Surface-PEGylated betaCDP polyplexes are formed by self-assembly of the polyplexes with adamantane-PEG conjugates. While unmodified polyplexes rapidly aggregate and precipitate in salt solutions, the PEGylated betaCDP polyplexes are stable at conditions of physiological salt concentration. Addition of targeting ligands to the adamantane-PEG conjugates allows for receptor-mediated delivery; galactosylated betaCDP-based particles reveal selective targeting to hepatocytes via the asialoglycoprotein receptor. Galactosylated particles transfect hepatoma cells with 10-fold higher efficiency than glucosylated particles (control), but show no preferential transfection in a cell line lacking the asialoglycoprotein receptor. Thus, surface modification of betaCDP-based polyplexes through the use of cyclodextrin/adamantane host/guest interactions endows the particles with properties appropriate for systemic application.

Adamantane↗

Temperature-actuated changes in wettability at elastomer/water interfaces.

Surface modification of 1,4-polybutadiene and cis-1,4-polyisoprene to introduce polar functional groups provided surfaces that reconstructed reversibly against water as a function of temperature. These surfaces became hydrophobic in contact with hot water, but their original hydrophilicity returned upon equilibration against cold water. Repeated cycling between hot and cold water, however, led to a damping of this reversibility. A series of parallel experiments on both the interfacial and bulk behavior of these elastomers strongly indicated that this damping was due to the alignment of extended interfacial chains during temperature cycling and to a decay of the restoring force on the interfacial chains under extension. These studies thus demonstrate that the interfacial behavior of elastomers can display close analogies to the bulk viscoelastic properties of the solid.

Journal Article↗

Glutaraldehyde modification of recombinant human hemoglobin alters its hemodynamic properties.

Many cell-free hemoglobin solutions designed as oxygen-carrying therapeutics produce a hypertensive effect in animals. The response is likely due to oxidation of nitric oxide by hemoglobin. Since the site of oxidation may lie outside the vascular compartment, we tested the hypothesis that polymerization of hemoglobin, rHb1.1, by glutaraldehyde would attenuate the hypertensive response. Two products of the cross-linking reaction were isolated, a glutaraldehyde-derivatized monomer (mono-glxrHb) and a glutaraldehyde cross-linked polymer (poly-glxrHb), and evaluated for their effects on systemic hemodynamics in conscious rats. Administration of rHb1.1 caused a mean arterial pressure elevation of approximately 20 mm Hg and an increase in total peripheral resistance of approximately 30%. Administration of mono-glxrHb induced changes in mean arterial pressure and vascular resistance that were significantly diminished relative to those observed with rHb1.1. Poly-glxrHb elicited a mean arterial pressure response that was further reduced compared with that obtained with mono-glxrHb and a change in vascular resistance that was the same as the response to mono-glxrHb. These results suggest that rHb peripheral vasoconstriction elicited by rHb1.1 is significantly attenuated by glutaraldehyde modification of the hemoglobin monomer and that the effect of glutaraldehyde polymerization is likely due to surface modification and/or intramolecular cross-linking, rather than an increase in molecular size.

Animals↗

The biomechanical and histopathologic effects of surface texturing with silicone and polyurethane in tissue implantation and expansion.

There has been considerable interest in determining the effect of morphologic alterations of prosthetic surfaces on capsule response in breast surgery. The purpose of this study was to provide a precise, three-dimensional evaluation of soft-tissue response to surface modifications in both implantation and expansion. Expandable 100-cc prostheses were designed with one of three surfaces: textured silicone (Biocell), standard smooth silicone, or polyurethane (Natural-Y, Meme). A new submuscular implantation site in the rabbit was developed. Each animal randomly received a smooth-surface device on one side and either a textured silicone or polyurethane device on the other. In one group of animals, the prostheses were expanded monthly. Capsular response was evaluated monthly in vivo using standardized techniques as well as biomechanical methods for up to 6 months in the expander group (n = 7 to 16) and 8 months in the implant group (n = 7 to 15). Analysis of biomechanical and histologic data revealed that prosthetic surface morphology can specifically alter capsular response. Polyurethane was the only effective surface in preventing capsular contracture in implantation. In expansion, both textured silicone and polyurethane surfaces resulted in significantly less capsular contracture and less resistance to expansion than comparable smooth-surfaced controls. Statistical comparisons reveal that the biomechanical methods utilized in this study provide the most precise and objective method of defining overall soft-tissue contracture around implanted biomaterials.

Animals↗

Biomaterials: where we have been and where we are going.

Since its inception just over a half century ago, the field of biomaterials has seen a consistent growth with a steady introduction of new ideas and productive branches. This review describes where we have been, the state of the art today, and where we might be in 10 or 20 years. Herein, we highlight some of the latest advancements in biomaterials that aim to control biological responses and ultimately heal. This new generation of biomaterials includes surface modification of materials to overcome nonspecific protein adsorption in vivo, precision immobilization of signaling groups on surfaces, development of synthetic materials with controlled properties for drug and cell carriers, biologically inspired materials that mimic natural processes, and design of sophisticated three-dimensional (3-D) architectures to produce well-defined patterns for diagnostics, e.g., biological microelectromechanical systems (bioMEMs), and tissue engineering.

Adsorption↗

Correlated scanning and transmission electron microscopy of developing lens vesicle in the chick.

Cells within the lens placode undergo transient surface modifications during the course of invagination. At stage 13, cellular apices bulge outward and display decreased surface diameters when compared to adjacent surface ectoderm. The lens vesicle is well invaginated by stage 14. An area of modified cells can be recognized ventral to the lens vesicle aperture. These cells present a smaller surface area and increased pleomorphism. Normal surface ectoderm extends to the dorsal and lateral edges of the aperture. The apical surfaces of cells which line the lumen of the lens vesicle display a complex topography such that individual cell boundaries are not easily distinguished. At stage 17, cellular processes which partially occlude the closing aperture of the lens vesicle may assist in reestablishing ectoderm over the newly formed lens. Preliminary results of cytochalasin B exposure on the invaginating lens vesicle suggest that the contractile action of microfilaments influences the complex cellular topography.

Animals↗

Cementless implant fixation--toward improved reliability.

Cementless implants offer the advantage of fixation by direct bone-to-implant osseointegration, thereby avoiding the use of a synthetic intermediary material (such as acrylic bone cement) of limited mechanical strength. Successful osseointegration, however, depends on several conditions being satisfied during the peri-implant bone healing period, including the need for limited early loading resulting in minimal relative movement at the implant-bone interface. Sintered porous- and plasma spray-coated implants represent the most common cementless orthopedic implants in current clinical use, although novel cast structures also are being investigated. All stand to benefit from surface modifications currently being explored to enhance osteoconductive or osteoinductive characteristics of the implants. The faster osseointegration that such modified surface designs potentially might offer would result in more reliable and convenient (from the patient perspective) cementless implants. Encouraging results of early animal-based studies exploring such modifications have been reported.

Alloys↗

Two different pathways for necrotic cell death induced by free radicals.

Plasma membrane modifications have been widely recognized as crucial factors in cell injury and death. One of these modifications, surface blebbing, has been considered as an injury-marker associated with a series of biochemical and physiological modifications. Our study focused on the different effects of free radical-induced cell damage by quinone menadione (2-methyl-1,4-naphthoquinone) and by hyperthermic shock (45 degrees C) on the erythroleukemic cell line K562. Different techniques including immunofluorescence, freeze-fracturing, and electron paramagnetic resonance spectroscopy were employed. Menadione induced the formation of surface blebs, accompanied by a rearrangement of the microfilament system and changes in the distribution of plasma membrane proteins. In contrast, heat-shocked cells showed neither blebbing nor important cytoskeletal changes. Finally, the electron paramagnetic resonance results showed an increase in membrane order not specifically related to the type of free radical-induced stress. These cell death features appear to suggest the existence of two different types of pathways for necrotic cell death: both treatments induce cell injury and eventual death by modifying plasma membrane integrity and function. However, one involves cytoskeleton-dependent surface blebbing, whereas the other does not.

Cell Death↗

Polymers for biodegradable medical devices. VII. Hydroxybutyrate-hydroxyvalerate copolymers: degradation of copolymers and their blends with polysaccharides under in vitro physiological conditions.

The hydrolytic degradation of hydroxybutyrate-hydroxyvalerate copolymers was monitored in vitro at 37 degrees C and pH 7.4. Direct use of bulk properties such as weight loss and tensile strength did not reveal substantial changes in the polymer matrix over degradation periods of several months. Despite this, the polymers were demonstrated to undergo significant modification during this period, in ways that markedly influence their subsequent behaviour. Combined use of goniophotometry and surface energy measurements revealed that surface modification begins at an early stage and is accompanied by diffusion of water into the matrix and a progressive increase in polymer porosity. Relatively little change in the molecular weight and some increase in the crystallinity of the matrix occurred during these early months. As a result, the tensile strength of the polymer varies little in this period. As the porosity of the matrix increases, hydrolytic chain scission within the matrix and diffusion out of degradation products proceeds more effectively. Decrease in matrix molecular weight, increase in matrix erosion, weight loss and loss of tensile strength began at a much more dramatic rate. The apparent resistance of the polymer to degradation in the early months is followed by an accelerated degradation phase around and beyond 1 yr. The use of filters that can dissolve or hydrolytically degrade more rapidly than the hydroxybutyrate matrix accelerates the development of porosity within the matrix and thus enhances the decomposition process.

Biocompatible Materials↗

Surface immobilization of galactose onto aliphatic biodegradable polymers for hepatocyte culture.

A novel surface modification method of biodegradable polymers was investigated for inducing the attachment of specific cells onto the polymer surface via ligand-receptor interactions. Galactose, a targeting ligand specific to asialoglycoprotein receptors present on cell membrane of hepatocytes, was introduced on the surface of poly(D,L-lactic-co-glycolic acid) (PLGA) films. A terminal end group of carboxylic acid in PLGA was activated by dicyclohexylcarbodiimide and N-hydroxysuccinimide for the direct conjugation of lactose by reductive amination reaction. Di-block copolymers of PLGA-b-poly(ethylene glycol) (PEG) having a free terminal amine group were also synthesized and used for the conjugation of galactose for the introduction of a PEG spacer between PLGA and galactose. The presence of galactose moieties on the blend film surface was characterized by measuring water contact angle and X-ray photon spectroscopy, and the amount of galactose was indirectly determined by a specific lectin-binding assay. With increasing the galactose concentration on the blend film surface, the initial attachment as well as the cell viability of hepatocyates concomitantly increased. The introduction of PEG spacer reduced the cell attachment and viability. Albumin secretion rate from hepatocytes was enhanced for galactose modified surfaces, whereas it was reduced for the surfaces not having galactose moieties.

Animals↗

Reactions of a Trifunctional Silane Coupling Agent in the Presence of Colloidal Silica Sols in Polar Media.

Reactions of 3-glycidoxypropyltrimethoxysilane (GPS) in a water-rich environment with silica colloids were studied. (29)Si NMR spectroscopy was used to monitor quantitatively the variation in GPS hydrolysis, condensation, and adsorption with pH and water concentration. The results show the competition between adsorption on the silica surface and condensation polymerization of GPS in the bulk liquid and are used to identify conditions favorable to surface modification. The presence of colloidal silica was shown to increase the apparent GPS dimerization rate, suggesting either surface catalysis or an increased local reactant concentration in the electrical double layer around the silica colloids. Copyright 1999 Academic Press.

Journal Article↗

[Surface characterization of Al2O3-blasted titanium implants in total hip arthroplasty].

INTRODUCTION: Shot peening and grit blasting techniques are used in several surface modifications for producing hip endoprostheses. Corund blasting techniques using highly pure Al2O3 particles to create a rough surface of titanium implants in cementless total hip arthroplasty achieve better osteointegration. An increasing number of publications in maxillofacial surgery and orthopedic surgery show that there is a significant contamination on corund-blasted surfaces. The latest research studies reported an effect of contaminant particles on early failure of endoprostheses associated with third body wear. The aim of our work was to evaluate the amount and particle size of Al2O3 on the implant surface. MATERIAL AND METHOD: The surface of four different geometries (five parts each, ARR titanium acetabular reinforcement ring, anisotropic Vektor cup, Vektor titanium stem and modular MRP Titan stem) were analyzed with respect to Al2O3 particles. Grit blasting was performed with Al2O3 particles, Alodur SWSK, Fa. Treibacher. Field emission scanning electron microscopy (LEO 1525) was used for the detection of the Al2O3 particles at the implant surface with a backscattered electron detector. The particle size distribution as well as the average area of the covered surface was calculated with image analyzing software (analySIS, Soft Imaging System GmbH). RESULTS: The surface of the anisotropic Vektor cup was contaminated at an average of 41.7+/-4.9%, the Vektor titanium stem at an average of 33.3+/-4.7%, the (MRP) Titan stem at an average of 30.6+/-4.2%, and the ARR titanium acetabular reinforcement ring at an average of 23.2+/-1.6 with Al2O3 particles over the whole surface orientated to the bone. DISCUSSION: The results of this study clearly show that there is contamination of rough titanium surfaces with Al2O3 particles at an unexpectedly high percentage. With respect to third body wear in total hip arthroplasty, further studies are necessary to minimize contamination of roughened surfaces and maintain sufficient roughness for osteointegration.

Aluminum Oxide↗

Protein bonding on biodegradable poly(L-lactide-co-caprolactone) membrane for esophageal tissue engineering.

A biodegradable and flexible poly(L-lactide-co-caprolactone) (PLLC) copolymer was synthesized and surface modification has been performed aiming at application as a scaffold in esophageal tissue engineering. The PLLC membrane surface was aminolyzed by 1,6-hexanediamine to introduce free amino groups. Using these amino groups as bridges, fibronectin and collagen were subsequently bonded with glutaraldehyde as a coupling agent. The presence of free amino groups on the aminolyzed PLLC surface was quantified using fluorescamine analysis method, which revealed that the surface NH2 density increased and eventually saturated with increasing 1,6-hexanediamine concentration or reaction time. X-ray photoelectron spectroscopy (XPS) confirmed the presence of both proteins separately on the modified PLLC surface. Water contact angle measurements evaluate the wettability of modified and unmodified PLLC surfaces. Protein-bonded surface presented more hydrophilic and homogeneous, yet PLLC can also adsorb some protein molecules. In vitro long-term (12d) culture of porcine esophageal cells proved that fibronectin- and collagen-modified PLLC surface (denoted PLLC-Fn and PLLC-Col, respectively) can more effectively support the growth of smooth muscle cells and epithelial cells; both modified and unmodified PLLC support fibroblasts growth. Mitochondrial activity assay and cell morphology observation indicate that the PLLC-Fn surface is more favorable to epithelium regeneration than PLLC-Col. These culture results provide much valuable information for our subsequent research on the construction of artificial scaffolds with esophageal function. Fibronectin-integrated PLLC will be a good candidate scaffold to support the growth of all types of esophageal cells.

Acetamides↗

A biochip platform for cell transfection assays.

In this paper, we describe the development and characterization of a biochip platform for cell transfection assays. Silicon wafers were surface modified by plasma polymerization of allylamine plasma polymer (ALAPP) and grafting of a protein-resistant layer of poly(ethylene oxide) (PEO) on the plasma polymer surface. Excimer laser ablation was then used to pattern ALAPP-PEO coated samples for spatially controlled protein adsorption and subsequent cell attachment. X-ray photoelectron spectroscopy (XPS) was used to characterize the surface modifications before and after excimer laser ablation. Experiments confirmed the creation of a two-dimensionally controlled surface chemistry on the biochip. Cell culture experiments using human embryonic kidney (HEK 293) cells showed that cells attached exclusively to laser ablated areas. In addition, cells confined to ablated areas were successfully transfected with plasmid DNA containing the gene for green fluorescent protein (GFP). The cell transfection efficiencies of cells growing in a culture flask and cells confined on the biochip were determined to be 21 and 13%, respectively.

Biological Assay↗