Biomaterials (v.26, #28)

Calendar (I).

Effect of cobalt and chromium ions on MMP-1, TIMP-1, and TNF-α gene expression in human U937 macrophages: A role for tyrosine kinases by Li Luo; Alain Petit; John Antoniou; David J. Zukor; Olga L. Huk; Roger C.W. Liu; Françoise M. Winnik; Fackson Mwale (5587-5593).
Previous reports have suggested that the imbalance of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) activity may contribute to prosthetic loosening. However, the mechanisms controlling these enzymes in the periprosthetic environment is unknown. We examined the effect of Co2+ and Cr3+ ions on the expression of genes encoding MMP-1, one of the principal proteinases capable of degrading native fibrillar collagens in the extracellular matrix (ECM), its inhibitor TIMP-1, and TNF-α, a cytokine that plays a central role in the induction of implant osteolysis. Human U937 macrophages were incubated in suspension or on phosphorylcholine (PC)-polymer coated surfaces for 24 h with Co2+ and Cr3+ ions. The level of mRNAs was determined by reverse transcription-polymerase chain reaction (RT–PCR). Results show that both Co2+ and Cr3+ ions induce the expression of MMP-1, TIMP-1, and TNF-α mRNA in a dose-dependent manner in cell suspensions. Tyrosine kinase inhibitors have different effects on these stimulatory effects. Indeed, genistein has only partial inhibitory effect on MMP-1 and TIMP-1, with even less effect on TNF-α expression. In contrast, herbimycin A completely blocks MMP-1 and TNF-α while partially inhibiting TIMP-1. However, Co2+ and Cr3+ ions had no effect on the expression of MMP-1 and TIMP-1 in macrophages cultured on the PC-polymer, suggesting that the attachment of U937 macrophages to the PC-polymer surfaces may modify their gene expression. In fact, MMP-1 and TIMP-1 seems to be constitutively up-regulated in this condition. However, the effect of Co2+ and Cr3+ ions on macrophages cultured on PC-polymer coated surfaces is similar to what was observed in suspension. Together, these findings indicate that activation of MMP-1, TIMP-1, and TNF-α by Co2+ and Cr3+ ions is regulated by tyrosine kinases.
Keywords: Cell adhesion; Hip replacement prosthesis; Macrophage; Matrix metalloproteinase; Metal ions; Phosphorylcholine;

Effect of vitamin E addition to poly(d,l)-lactic acid on surface properties and osteoblast behaviour by Filippo Renò; Valentina Aina; Silvia Gatti; Mario Cannas (5594-5599).
Vitamin E (Vit.E, α −tocoferol) is a natural agent with anti-oxidative and anti-inflammatory properties and it has been suggested that it could act as a stimulating factor for osteoblast proliferation and maturation. We produced poly(d,l)-lactic acid films enriched with Vit.E (1, 5 and 10% w/w) and investigated their surface properties using the FTIR analysis, sessile measure of wettability and serum protein adsorption, and evaluated attachment and spreading of MC-3T3 E1 murine osteoblast cells. FTIR analysis showed the presence of Vit.E on the polymer surface and Vit.E increased the polymer wettability in a concentration-dependent manner.The serum total protein adsorption increased significantly onto the 10% Vit.E P(d,l)-LA and the main protein adsorbed was albumin. The presence of albumin, considered as an anti-adhesive protein, on the surface of Vit.E enriched P(d,l)-LA films (especially 5 and 10% Vit.E) could explain, at least in part, the behaviour of MC-3T3 osteoblast cells seeded onto the polymers. Cell adhesion and spreading were strongly decreased by Vit.E (5 and 10%) in spite of the increased wettability. This reaction could be cell type-specific, independent by the surface wettability and linked to cell-specific characteristics (membrane phospholipid composition, integrins expression). Moreover a direct effect of Vit.E on cell adhesion and spreading cannot be completely excluded.
Keywords: P(d,l)-LA; Vitamin E; Osteoblast;

Bone formation and resorption of highly purified β-tricalcium phosphate in the rat femoral condyle by Naoki Kondo; Akira Ogose; Kunihiko Tokunaga; Tomoyuki Ito; Katsumitsu Arai; Naoko Kudo; Hikaru Inoue; Hiroyuki Irie; Naoto Endo (5600-5608).
The aim of this study was to examine the chronological histology associated with highly purified β-tricalcium phosphate (β-TCP) implanted in the rat femoral condyle. Specimens were harvested on days 4, 7, 14, 28 and 56 after implantation, and were analyzed by tartrate-resistant acid phosphatase (TRAP) staining, immunohistochemistry of the ED1 protein as a marker of the phagocyte system, and in situ hybridization with digoxigenin-labeled α1 chain of type I procollagen (COL1A1), osteopontin and osteocalcin. β-TCP was resorbed in a chronological manner. Although new bone was not observed on day 4, fibroblast-like cells around β-TCP were positive for COL1A1 and osteopontin mRNA. New bone formation presented after day 7. In the double-staining for OPN and ED1 on day 7, most cells around β-TCP were positive for either osteopontin mRNA or ED1 protein. However, there were some doubly positive multinucleated cells, suggesting that they belonged to the mononuclear phagocyte system. After day 28, the implanted region was replaced with bone marrow. Multinucleated TRAP-positive and ED1-positive cells which adhered to β-TCP at all stages seemed to be osteoclasts and they continuously resorbed β-TCP. β-TCP has a good biocompatibility since both bioresorption and bone formation started at an early stage after implantation.
Keywords: Bioresorption; Calcium phosphate; Macrophage; Osteoblast; Osteoclast;

Developing macroporous bicontinuous materials as scaffolds for tissue engineering by Monique Martina; Gayathri Subramanyam; James C. Weaver; Dietmar W. Hutmacher; Daniel E. Morse; Suresh Valiyaveettil (5609-5616).
Calcareous skeletal elements (ossicles) isolated from the seastar, Pisaster giganteus, were characterized and tested as potential biocompatible substrates for cellular attachment. These ossicles have a remarkably robust open-framework architecture with an interconnected network of ca. 10 μm diameter pores. Scanning electron and confocal microscopy was used to characterize the cell-substrate interaction. Cell culturing experiments revealed that the cells firmly attach to the ossicle surface, forming cell aggregates of several layers thick. The anchored cells extended to form ‘bridges’ between the openings in the bicontinuous framework and the degree of coverage increased as culture time progressed. Osteoblasts grown on the ossicles were found to be viable up to 32 days after initial seeding, as proven by assaying with AlamarBlue™ and FDA/PI staining indicating the ossicle's potential as an alternative highly effective tissue scaffold. Given the limitation in availability of this natural material, the results presented here should be seen as offering guidelines for future development of synthetic materials with physical and chemical properties strongly conducive to bone repair and restoration.
Keywords: Bone substitutes; Cell adhesion; Cell morphology; Bone graft; Osteoblast;

Preparation of γ -PGA/chitosan composite tissue engineering matrices by Chien-Yang Hsieh; Sung-Pei Tsai; Da-Ming Wang; Yaw-Nan Chang; Hsyue-Jen Hsieh (5617-5623).
γ -poly(glutamic acid) ( γ -PGA), a hydrophilic and biodegradable polymer, was chosen to modify chitosan matrices to produce a γ -PGA/chitosan composite biomaterial. Three types of both dense and porous composite matrices containing different amounts of γ -PGA were fabricated. Chitosan and γ -PGA matrices were also prepared as controls. Fluorescence staining indicated that chitosan and γ -PGA were evenly distributed in the composite matrices. SEM micrographs showed that an interconnected porous structure with a pore size of 30–100 μm was present in all porous matrices except the γ -PGA ones. By increasing the percentage of γ -PGA from 0% to 20%, the swelling ratio of the matrices was enhanced from 1.6 to 3.2. Similarly, the contact angle of the matrices decreased from 113° to 94°. These data suggested that the surface hydrophilicity, water absorption rate, and swelling ratio were improved by adding γ -PGA to the matrices. Additionally, the mechanical strength of the porous γ -PGA/chitosan matrices was about 25–50%, higher than that of the unmodified chitosan matrices. The composite matrices were also examined and found to be an appropriate environment for cell attachment and proliferation. The cell density on the 20% γ -PGA-modified matrices was almost triple that on the unmodified chitosan matrices on day 5. In summary, the γ -PGA/chitosan composite matrices, due to their better hydrophilic, cytocompatible, and mechanical properties, are very promising biomaterials for tissue engineering applications.
Keywords: Chitosan; γ -PGA; Tissue engineering; Composite matrices;

A synthetic nanofibrillar matrix promotes in vivo-like organization and morphogenesis for cells in culture by Melvin Schindler; Ijaz Ahmed; Jabeen Kamal; Alam Nur-E-Kamal; Timothy H. Grafe; H. Young Chung; Sally Meiners (5624-5631).
The purpose of this study was to design a synthetic nanofibrillar matrix that more accurately models the porosity and fibrillar geometry of cell attachment surfaces in tissues. The synthetic nanofibrillar matrices are composed of nanofibers prepared by electrospinning a polymer solution of polyamide onto glass coverslips. Scanning electron and atomic force microscopy showed that the nanofibers were organized into fibrillar networks reminiscent of the architecture of basement membrane, a structurally compact form of the extracellular matrix (ECM). NIH 3T3 fibroblasts and normal rat kidney (NRK) cells, when grown on nanofibers in the presence of serum, displayed the morphology and characteristics of their counterparts in vivo. Breast epithelial cells underwent morphogenesis to form multicellular spheroids containing lumens. Hence the synthetic nanofibrillar matrix described herein provides a physically and chemically stable three-dimensional surface for ex vivo growth of cells. Nanofiber-based synthetic matrices could have considerable value for applications in tissue engineering, cell-based therapies, and studies of cell/tissue function and pathology.
Keywords: Biomimetic material; Nanotopography; ECM; Cell culture;

Concentrated hydroxyapatite inks for direct-write assembly of 3-D periodic scaffolds by Sarah Michna; Willie Wu; Jennifer A. Lewis (5632-5639).
Hydroxyapatite (HA) scaffolds with a 3-D periodic architecture and multiscale porosity have been fabricated by direct-write assembly. Concentrated HA inks with tailored viscoelastic properties were developed to enable the construction of complex 3-D architectures comprised of self-supporting cylindrical rods in a layer-by-layer patterning sequence. By controlling their lattice constant and sintering conditions, 3-D periodic HA scaffolds were produced with a bimodal pore size distribution. Mercury intrusion porosimetry (MIP) was used to determine the characteristic pore size and volume associated with the interconnected pore channels between HA rods and the finer pores within the partially sintered HA rods.
Keywords: Hydroxyapatite; Bone tissue engineering; Scaffolds; Direct-write assembly;

Subcutaneous xenotransplantation of bovine pancreatic islets by Marina Figliuzzi; Roberta Cornolti; Tiziana Plati; Navneeta Rajan; Federica Adobati; Giuseppe Remuzzi; Andrea Remuzzi (5640-5647).
Transplantation of pancreatic islets in diabetes is currently limited by the need of immunosuppressive therapy. The present study was designed to test an immunoprotection planar device for subcutaneous xenotransplantation of pancreatic islets in the diabetic rat. We tested three different devices made of polyethersulfone hollow fibers. In all diabetic rats, implantation of islet-containing devices promptly normalized hyperglycemia. In vitro membrane permeability to glucose was correlated with implant function duration. These data confirm that bovine islets contained within devices and implanted subcutaneously remain functional for several days. Strategies to prolong islet function may allow achieving successful long-term islet implantation in this attractive site.
Keywords: Pancreatic islets; Immunoisolation device; Hollow fibers; Subcutaneous site; Xenotransplantation;

Bioactivation of an anorganic bone matrix by P-15 peptide for the promotion of early bone formation by Michael Thorwarth; Stefan Schultze-Mosgau; Falk Wehrhan; Peter Kessler; Safwan Srour; Jörg Wiltfang; Karl Andreas Schlegel (5648-5657).
This animal experiment compared the regenerative processes within defined bony defects of the porcine skull after delivery of routinely utilized bone graft materials: anorganic bone matrix (ABM) and an identical ABM carrying the cell binding peptide P-15. Particulated autogenous bone was used as a control group. The chosen porcine model guaranteed the transferability of the obtained results to clinical practice. A total observation period of 6 months was defined. The bone samples were examined microradiographically and histologically at 8 specific times. Sufficient osseointegration and osseoconduction could be demonstrated for both anorganic bone minerals. However, in the selected model significantly higher mineralization rates ( p = 0.0286 ) were found in the microradiographic image at 12 weeks after application of the bioactive form. The histological examination confirmed this accelerating effect on bone formation starting at day 3. At the end of the study after 6 months, the mineralization values had equalized in both study groups.For the first time, the material was demonstrated to be suitable as a bone substitute material for the treatment of larger bony defects in a large animal model. The P-15 sequence accelerated the process of bone formation on the surface of the anorganic bone matrix as early as 3 days but was not traced over the whole term of the study.
Keywords: Bone regeneration; Anorganic bone matrix; Hydroxyapatite ceramics; P-15 peptide; Bone graft; Collagen;

Cells can sense the physical and chemical properties of artificial materials used as scaffolds for tissue engineering and regulate their behavior. Therefore, biomimetic and biospecific molecules are coated on materials to regulate function of cells on the tissue-engineered product. These bioactive molecules can be attached in a defined spectrum, concentration and spatial distribution in order to control adhesion, growth, viability, differentiation, and function of the cells. When autologous cells are used for tissue engineering, initially limited cells obtained may often need an amplification of cell number by passage in tissue culture before they are seeded on a biomaterial or scaffold. We have conducted this study to understand how the characteristics of bioactive molecule coating might affect proliferation, apoptosis and differentiation when endothelial cell (EC) is serially passaged. Proliferation was assessed by proliferating cell nuclear antigen (PCNA) staining along with counting of cells harvested from confluent monolayer. Apoptosis was assessed by Annexin V staining and differentiation by semi quantitative reverse transcriptase polymerase chain reaction (RT-PCR) for von Willebrand factor (vWF) expression and quantification of its release using enzyme linked immunosorbant assay (ELISA), and thrombogenicity by comparing platelet adhesion to EC monolayer Dacron grafts (DG) with specific protein coating. The results indicate that ECs easily lose its proliferation potential when they are cultured repeatedly on gelatin, turn apoptotic and over express the prothrombotic protein- vWF. Whereas, when it is grown on a matrix composed of fibrin, fibronectin, gelatin and vascular EC growth factor (VEGF), the cells retained their ability to proliferate, remained viable and were relatively less thrombogenic, even when passage number progressed. It is concluded that if ECs are grown on the composite matrix that mimics natural vessel scaffold, the cell number can be amplified without affecting its normal physiological function and may be used to generate effective tissue-engineered cardiovascular constructs.
Keywords: Endothelial Cell; Vascular graft; Extracellular matrix; FIBRIN; Gelatin; Phenotypic alteration;

Down regulation of degenerative cartilage molecules in chondrocytes grown on a hyaluronan-based scaffold by Brunella Grigolo; Luciana De Franceschi; Livia Roseti; Luca Cattini; Andrea Facchini (5668-5676).
Hyaluronic-acid-based biomaterials used for cartilage repair allow the expression of specific extracellular matrix molecules by human chondrocytes grown onto them. We investigated whether these biomaterials could also create an environment in which the cells downregulate the expression of some catabolic factors.Chondrocytes were isolated from human articular cartilage obtained from the knees of patients with a history of trauma. First, the cells were expanded in monolayers and then they were seeded on a hyaluronic-acid derivative scaffold. Constructs and surnatants were collected and analysed at 1, 3, 7, 14 and 21 days after seeding. Immunohistochemical analysis for CD44 and caspase was carried out on paraffin-embedded sections. The Tunel method was used to identify chondrocyte apoptosis status. Secretion of MMP-1 and MMP-13 in the surnatants of the cells grown onto the biomaterial was measured by enzyme-linked immunosorbent assay. Nitric oxide (NO) production was evaluated by estimating the stable NO metabolite nitrite by the Griess method. A real-time RT-PCR analysis was performed on the constructs to evaluate the expression of type I and II collagens, aggrecan, Sox-9, MMP-1 and MMP-13 mRNAs at the different experimental times evaluated.Decreased levels of metalloproteinases and nitric oxide were observed in the surnatants of chondrocytes grown onto the hyaluronan-based scaffold. This was also confirmed by real-time PCR analysis which showed that the cells expressed the specific differentiated phenotype downregulating the expression of some catabolic molecules. Cells apoptosis decreased during the culture period, which further supported the biochemical data.The ability of the hyaluronan scaffold to reduce the expression and production of molecules involved in cartilage degenerative diseases indicates its use to treat early lesions of osteoarthritic patients.
Keywords: Chondrocytes; Hyaluronic acid; Arthritis; Apoptosis; Metalloproteinases;

Cellular and molecular events during chondrogenesis of human mesenchymal stromal cells grown in a three-dimensional hyaluronan based scaffold by Gina Lisignoli; Sandra Cristino; Anna Piacentini; Stefania Toneguzzi; Francesco Grassi; Carola Cavallo; Nicoletta Zini; Liliana Solimando; Nadir Mario Maraldi; Andrea Facchini (5677-5686).
Mesenchymal stromal cells (MSCs) seem to be a good alternative to chondrocytes for cartilage regeneration. To obtain new information on the sequence of cellular and molecular events during in vitro chondrogenic differentiation we analysed MSCs on a widely used hyaluronic acid biomaterial (Hyaff®-11). Cellular differentiation was induced using two different concentrations of TGF β 1 (10 and 20 ng/ml) and the process was analysed at different time points (24 h, and 7, 14, 21 and 28 days) using techniques of light and electron microscopy, real-time PCR and immunohistochemistry. We found that without TGF β MSCs did not survive while in the presence of TGF β the cells significantly proliferated from day 7 until day 28. Light and electron microscopy showed that TGF β at 20 ng/ml better induced the formation of cartilage-like tissue. Real-time PCR showed an increased expression of collagen type II, IX and aggrecan associated to a down-regulation of collagen type I. Immunohistochemical analysis confirmed that collagen type I was down-modulated while collagen type II increased from day 14 to day 28.These data clearly showed that higher concentrations of TGF β (20 ng/ml) induce chondrogenesis of MSCs on Hyaff®-11 scaffold better than 10 ng/ml of TGF β . This process is characterized by a sequence of cellular and molecular events that deal with the in vitro formation of a cartilage-like tissue.
Keywords: Mesenchymal stromal cells; Chondrogenesis; Biomaterial; Tissue engineering;

Effect of bone mineral density and amorphous diamond coatings on insertion torque of bone screws by Arto Koistinen; Seppo S. Santavirta; Heikki Kröger; Reijo Lappalainen (5687-5694).
In this study, the potential of high-quality amorphous diamond (AD) coatings in reducing the torque and failures of bone screws was studied. Torque values were recorded for 32 stainless steel screws, 2.7 or 3.5 mm in diameter and 60 mm in length. Half of the screw sets were coated with the AD coating before installing in predrilled holes of human cadaveric femoral bone samples. The bone samples were selected from two groups of four persons with mean ages of 34 years (range 25–41 years) and 75 years (range 73–77 years), respectively. The bone mineral density (BMD) values of the samples were determined exactly at the screw insertion site by peripheral quantitative computed tomography (pQCT). In the mechanical tests, insertion and removal torques were measured. BMD had a significant effect on insertion torque; the maximum torque (adjusted with respect to the screw diameter) was significantly higher for the young bone than for the old bone ( p < 0.05 ). By using a polished AD coating, insertion torque was decreased even up to 50% in comparison with the screws without coating. The results suggest that AD coating provides a stable, smooth surface and reduces the risk of screw failures.
Keywords: Biocompatibility; Bone repair; Diamond; Mechanical test; Surface treatment;

Fe-based nanoparticulate metallic alloys as contrast agents for magnetic resonance imaging by Oscar Bomatí-Miguel; María P. Morales; Pedro Tartaj; Jesús Ruiz-Cabello; Pierre Bonville; Martín Santos; Xinqing Zhao; Sabino Veintemillas-Verdaguer (5695-5703).
Pharmaceutical grade magnetic colloidal dispersions have been prepared from iron alloys synthesized by laser pyrolysis. The colloids were obtained by simultaneous dispersion and coating of the particles with dextran in a strong alkaline solution. Both powders and dispersions have been analyzed in terms of microstructural characteristics, chemical composition and magnetic properties. The powders consist of uniform spherical nanoparticles (12 nm of diameter) showing a metallic core encapsulated into an iron-oxide shell. On the other hand, the colloidal dispersions consist of magnetic particles-aggregates with hydrodynamic sizes of ∼75 nm. Magnetic resonance images of rats were taken after the intravenously administration of the Fe colloidal dispersions, and compared with those obtained using a commercial iron oxide magnetic resonance imaging contrast agent. The results showed a contrast improvement of 60% in the liver with respect to the commercial sample, which suggests that this product could be a suitable contrast agent for NMR imaging of liver and spleen.
Keywords: Laser; Nanoparticles; Magnetism; Magnetic resonance imaging (MRI); Liver;

Steady-state diffusion of water through soft-contact-lens materials by Francesco Fornasiero; Florian Krull; John M. Prausnitz; Clayton J. Radke (5704-5716).
Water transport through soft contact lenses (SCL) is important for acceptable performance on the human eye. Chemical-potential gradient-driven diffusion rates of water through SCL materials are measured with an evaporation-cell technique. Water is evaporated from the bottom surface of a lens membrane by impinging air at controlled flow rate and humidity. The resulting weight loss of a water reservoir covering the top surface of the contact-lens material is recorded as a function of time.New results are reported for a conventional hydrogel material (SofLens™ One Day, hilafilcon A, water content at saturation w 10 = 70 weight %) and a silicone hydrogel material (PureVision™, balafilcon A, w 10 = 36 % ), with and without surface oxygen plasma treatment. Also, previously reported data for a conventional 2-hydroxyethyl methacrylate (HEMA)-SCL ( w 10 = 38 % ) hydrogel are reexamined and compared with those for SofLens™ One Day and PureVision™ hydrogels. Measured steady-state water fluxes are largest for SofLens™ One Day, followed by PureVision™ and HEMA. In some cases, the measured steady-state water fluxes increase with rising relative air humidity. This increase, due to an apparent mass-transfer resistance at the surface (trapping skinning), is associated with formation of a glassy skin at the air/membrane interface when the relative humidity is below 55–75%.Steady-state water fluxes are interpreted through an extended Maxwell–Stefan diffusion model for a mixture of species starkly different in size. Thermodynamic nonideality is considered through Flory–Rehner polymer-solution theory. Shrinking/swelling is self-consistently modeled by conservation of the total polymer mass. Fitted Maxwell–Stefan diffusivities increase significantly with water concentration in the contact lens.
Keywords: Evaporation-cell method; Water diffusion-coefficient; Soft contact lens; Extended Maxwell–Stefan model; Trapping skinning; Glass transition;

The effect of the antimicrobial peptide, Dhvar-5, on gentamicin release from a polymethyl methacrylate bone cement by C. Faber; R.J.W. Hoogendoorn; D.M. Lyaruu; H.P. Stallmann; J. van Marle; A. van Nieuw Amerongen; T.H. Smit; P.I.J.M. Wuisman (5717-5726).
The objective of this study was to investigate the release mechanism and kinetics of the antimicrobial peptide, Dhvar-5, both alone and in combination with gentamicin, from a standard commercial polymethyl methacrylate (PMMA) bone cement. Different amounts of Dhvar-5 were mixed with the bone cement powders of Osteopal and the gentamicin-containing Osteopal G bone cement and their release kinetics from the polymerized cement were investigated. Additionally, the internal structure of the bone cements were analysed by scanning electron microscopy (SEM) of the fracture surfaces. Secondly, porosity was investigated with the mercury intrusion method and related to the observed release profiles. In order to obtain an insight into the mechanical characteristics of the bone cement mixtures, the compressive strength of Osteopal and Osteopal G with Dhvar-5 was also investigated. The total Dhvar-5 release reached 96% in the 100 mg Dhvar-5/g Osteopal cement, whereas total gentamicin release from Osteopal G reached only 18%. Total gentamicin release increased significantly to 67% with the addition of 50 mg Dhvar-5/g, but the Dhvar-5 release was not influenced. SEM showed an increase of dissolved gentamicin crystals with the addition of Dhvar-5. The mercury intrusion results suggested an increase of small pores (<0.1 μm) with the addition of Dhvar-5. Compressive strength of Osteopal was reduced by the addition of Dhvar-5 and gentamicin, but still remained above the limit prescribed by the ISO standard for clinical bone cements. We therefore conclude that the antimicrobial peptide, Dhvar-5, was released in high amounts from PMMA bone cement. When used together with gentamicin sulphate, Dhvar-5 made the gentamicin crystals accessible for the release medium presumably through increased micro-porosity (<0.1 μm) resulting in a fourfold increase of gentamicin release.
Keywords: Polymethylmethacrylate; Drug release; Porosity; Gentamicin; Dhvar-5; Antimicrobial peptide;

Surface modification of biodegradable polyesters with fatty acid conjugates for improved drug targeting by Tarek M. Fahmy; Robert M. Samstein; Casey C. Harness; W. Mark Saltzman (5727-5736).
We describe a general method for incorporating target ligands into the surface of biocompatible polyester poly(lactic-co-glycolic acid) (PLGA) 50/50 materials using fatty acids. Avidin-fatty acid conjugates were prepared and efficiently incorporated into PLGA. Avidin was chosen as an adaptor protein to facilitate the attachment of a variety of biotinylated ligands. We show that fatty acid preferentially associates with the hydrophobic PLGA matrix, rather than the external aqueous environment, facilitating a prolonged presentation of avidin over several weeks. We successfully applied this approach in both microspheres encapsulating a model protein, bovine serum albumin, and PLGA scaffolds fabricated by a salt-leaching method. Because of its ease, generality and flexibility, this strategy promises widespread utility in modifying the surface of PLGA-based materials for applications in drug delivery and tissue engineering.
Keywords: Biodegradation; Biocompatibility; Surface modification; Lipid; Drug delivery; Microsphere; Scaffold;

Chitosan beads as molecularly imprinted polymer matrix for selective separation of proteins by Tian-Ying Guo; Yong-Qing Xia; Jin Wang; Mou-Dao Song; Bang-Hua Zhang (5737-5745).
Two kinds of molecularly imprinted polymers were prepared using hemoglobin as the imprinting molecule, acrylamide as the functional monomer, chitosan beads and maleic anhydride-modified chitosan beads as matrixes, respectively. Static adsorbing experimental results showed that an equal class of adsorption was formed in the imprinted polymers and the adsorption equilibrium constant and the maximum adsorption capacity were evaluated. Chromatographic characteristics showed that the column bedded with the hemoglobin imprinted beads could separate hemoglobin and bovine serum albumin effectively from their mixture, which indicates that the imprinted beads have very higher selectivity for hemoglobin than the non-imprinted with the same chemical composition.
Keywords: Chitosan beads; Polyacrylamide; Protein separation; Molecular imprinting;

Embryonic stem (ES) cells have the ability to replicate themselves and differentiate into various mature cells. Recently, dopaminergic neurons were efficiently induced from ES cells using mouse stromal cells (PA6 cells) as a feeder cell layer. This simple procedure seems to be very efficient to obtain dopamine-releasing cells for future clinical cell transplantation treatment of Parkinson's disease. In this study, we prepared stock solutions containing neural inducing factors (NIFs) by washing PA6 cells with phosphate-buffered saline containing heparin. ES cells grew successfully in culture media supplemented with 33 v/v% NIFs stock solution, and the rate of neural differentiation of ES cell progeny increased with increasing heparin concentration in the culture media. In addition, NIFs-immobilized surfaces were prepared by exposing polyethyleneimine-modified surfaces to NIFs stock solutions. The NIFs-immobilized culture dish effectively supported cell growth as the culture medium supplemented with NIFs stock did, but its induction effect to dopaminergic neurons from ES cells was much smaller than free NIFs. NIFs stock solutions have two different activities. One can stimulate cell growth and the other induces differentiation of ES cells to the neural fate when heparin existed. The former factors were effectively immobilized on the culture dish, but those that induce differentiation may not be. Further optimization is required.
Keywords: Embryonic stem cell; Neural inducing factors; PA6 cell; Heparin; Neural cell; Surface modification;