Dual local drug delivery of vancomycin and farnesol for mitigation of MRSA infection in vivo - a pilot study.

Surgical site infections after orthopaedic surgery using fracture fixation devices or endosseous implants create major surgical challenges with severe adverse effects, such as osteomyelitis. These infections are frequently caused by Staphylococcus aureus, often with high resistance to antibiotics, such as methicillin-resistant Staphylococcus aureus (MRSA). Due to the formation of impenetrable biofilms on implant surfaces, systemic antibiotic treatment has become exceedingly difficult. New solutions are pursued by combining several drugs using a controlled delivery system from specifically engineered implant surfaces. A sol-gel coating on titanium implants was previously developed with 20 wt % vancomycin and 30 wt % farnesol, with suppression of MRSA in vitro. The present study investigated the efficacy of sol-gel film coatings for controlled dual local delivery over 4 weeks utilising a rat infection model. The findings confirmed the viability of this new concept in vivo based on the differences observed between coatings containing vancomycin alone (SGV) and the dual-drug-containing coating with vancomycin and farnesol (SGVF). While both the SGVF and SGV coatings facilitated excellent preservation of the osseous microarchitecture, SGVF coating displayed a slightly higher potency for suppressing MRSA infiltration than SGV, in combination with a lower reactive bone remodelling activity, most likely by disturbing biofilm formation. The next step for advancing the concept of dual-drug delivery from sol-gel coatings to the clinic and confirming the promising effect of the SGVF coatings on reactive bone remodelling and suppressing MRSA infiltration is a study in a larger animal species with longer time points.

Tyrosine-derived polycarbonate-silica xerogel nanocomposites for controlled drug delivery.

Biodegradable polymer-ceramic composites offer significant potential advantages in biomedical applications where the properties of either polymers or ceramics alone are insufficient to meet performance requirements. Here we demonstrate the highly tunable mechanical and controlled drug delivery properties accessible with novel biodegradable nanocomposites prepared by non-covalent binding of silica xerogels and co-polymers of tyrosine-poly(ethylene glycol)-derived poly(ether carbonate). The Young's moduli of the nanocomposites exceed by factors of 5-20 times those of the co-polymers or of composites made with micron scale silica particles. Increasing the fraction of xerogel in the nanocomposites increases the glass transition temperature and the mechanical strength, but decreases the equilibrium water content, which are all indicative of strong non-covalent interfacial interactions between the co-polymers and the silica nanoparticles. Sustained, tunable controlled release of both hydrophilic and hydrophobic therapeutic agents from the nanocomposites is demonstrated with two clinically significant drugs, rifampicin and bupivacaine. Bupivacaine exhibits an initial small burst release followed by slow release over the 7 day test period. Rifampicin release fits the diffusion-controlled Higuchi model and the amount released exceeds the dosage required for treatment of clinically challenging infections. These nanocomposites are thus attractive biomaterials for applications such as wound dressings, tissue engineering substrates and stents.

Elastic membrane that undergoes mechanical deformation enhances osteoblast cellular attachment and proliferation.

The main objective of this paper was to investigate the effect of transmission of force on bone cells that were attached to a deformable membrane. We functionalized a silastic membrane that measured 0.005 inches thickness and coated it with an extra cellular matrix (ECM) protein, fibronectin (FN). MC3T3-E1 osteoblast-like cells were cultured on the functionalized FN-coated membrane after which cell attachment and proliferation were evaluated. We observed an immediate attachment and proliferation of the bone cells on the functionalized membrane coated with FN, after 24 hours. Upon application of a mechanical force to cells cultured on the functionalized silicone membrane in the form of a dynamic equibiaxial strain, 2% magnitude; at 1-Hz frequency for 2 h, the osteoblast cells elicited slightly elevated phalloidin fluorescence, suggesting that there was reorganization of the cytoskeleton. We concluded from this preliminary data obtained that the engineered surface transduced applied mechanical forces directly to the adherent osteoblast cells via integrin binding tripeptide receptors, present in the FN molecules, resulting in the enhanced cellular attachment and proliferation.

Effect of functional end groups of silane self-assembled monolayer surfaces on apatite formation, fibronectin adsorption and osteoblast cell function.

Bioactive glass (BG) can directly bond to living bone without fibrous tissue encapsulation. Key mechanistic steps of BG's activity are attributed to calcium phosphate formation, surface hydroxylation and fibronectin (FN) adsorption. In the present study, self-assembled monolayers (SAMs) of alkanesilanes with different surface chemistry (OH, NH(2) and COOH) were used as a model system to mimic BG's surface activity. Calcium phosphate (Ca-P) was formed on SAMs by immersion in a solution that simulates the electrolyte content of physiological fluids. FN adsorption kinetics and monolayer coverage was determined on SAMs with or without Ca-P coating. The surface roughness was also examined on these substrates before and after FN adsorption. The effects of FN-adsorbed, Ca-P-coated SAMs on the function of MC3T3-E1 were evaluated by cell growth, expression of alkaline phosphatase activity and actin cytoskeleton formation. We demonstrate that, although the FN monolayer coverage and the root mean square (rms) roughness are similar on --OH and --COOH terminated SAMs with or without Ca-P coating, higher levels of ALP activity, more actin cytoskeleton formation and more cell growth are obtained on --OH- and --COOH-terminated SAMs with Ca-P coating. In addition, although the FN monolayer coverage is higher on Ca-P-coated --NH(2)-terminated SAMs and SiO(x) surfaces, higher levels of ALP activity and more cell growth are obtained on Ca-P-coated --OH- and --COOH-terminated SAMs. Thus, with the same Ca-P coatings, different surface functional groups have different effects on the function of osteoblastic cells. These findings represent new insights into the mechanism of bioactivity of BG and thereby may lead to designing superior constructs for bone grafting.

Effect of rapidly resorbable bone substitute materials on the temporal expression of the osteoblastic phenotype in vitro.

Ideally, bioactive ceramics for use in alveolar ridge augmentation should possess the ability to activate bone formation and, thus, cause the differentiation of osteoprogenitor cells into osteoblasts at their surfaces. Therefore, in order to evaluate the osteogenic potential of novel bone substitute materials, it is important to examine their effect on osteoblastic differentiation. This study examines the effect of rapidly resorbable calcium-alkali-orthophosphates on osteoblastic phenotype expression and compares this behavior to that of beta-tricalcium phosphate (TCP) and bioactive glass 45S5. Test materials were three materials (denominated GB14, GB9, GB9/25) with a crystalline phase Ca(2)KNa(PO(4))(2) and with a small amorphous portion containing either magnesium potassium phosphate (GB14) or silica phosphate (GB9 and GB9/25, which also contains Ca(2)P(2)O(7)); and a material with a novel crystalline phase Ca(10)[K/Na](PO(4))(7) (material denominated 352i). SaOS-2 human bone cells were grown on the substrata for 3, 7, 14, and 21 days, counted, and probed for an array of osteogenic markers. GB9 had the greatest stimulatory effect on osteoblastic proliferation and differentiation, suggesting that this material possesses the highest potency to enhance osteogenesis. GB14 and 352i supported osteoblast differentiation to the same or a higher degree than TCP, whereas, similar to bioactive glass 45S5, GB9/25 displayed a greater stimulatory effect on osteoblastic phenotype expression, indicating that GB9/25 is also an excellent material for promoting osteogenesis.

Materials in particulate form for tissue engineering. 1. Basic concepts.

For biomedical applications, materials small in size are growing in importance. In an era where 'nano' is the new trend, micro- and nano-materials are in the forefront of developments. Materials in the particulate form aim to designate systems with a reduced size, such as micro- and nanoparticles. These systems can be produced starting from a diversity of materials, of which polymers are the most used. Similarly, a multitude of methods are used to produce particulate systems, and both materials and methods are critically reviewed here. Among the varied applications that materials in the particulate form can have, drug delivery systems are probably the most prominent, as these have been in the forefront of interest for biomedical applications. The basic concepts pertaining to drug delivery are summarized, and the role of polymers as drug delivery systems conclude this review.

Materials in particulate form for tissue engineering. 2. Applications in bone.

Materials in particulate form have been the subjects of intensive research in view of their use as drug delivery systems. While within this application there are still issues to be addressed, these systems are now being regarded as having a great potential for tissue engineering applications. Bone repair is a very demanding task, due to the specific characteristics of skeletal tissues, and the design of scaffolds for bone tissue engineering presents several difficulties. Materials in particulate form are now seen as a means of achieving higher control over parameters such as porosity, pore size, surface area and the mechanical properties of the scaffold. These materials also have the potential to incorporate biologically active molecules for release and to serve as carriers for cells. It is believed that the combination of these features would create a more efficient approach towards regeneration. This review focuses on the application of materials in particulate form for bone tissue engineering. A brief overview of bone biology and the healing process is also provided in order to place the application in its broader context. An original compilation of molecules with a documented role in bone tissue biology is listed, as they have the potential to be used in bone tissue engineering strategies. To sum up this review, examples of works addressing the above aspects are presented.

RGDS peptides immobilized on titanium alloy stimulate bone cell attachment, differentiation and confer resistance to apoptosis.

A major cause of implant failure in skeletal tissues is failure of osseointegration, often due to lack of adhesion of cells to the titanium (Ti) alloy interface. Since arginine-glycine-aspartic acid (RGD)-containing peptides have been shown to regulate osteoblast adhesion, we tested the hypothesis that, bound to a Ti surface, these peptides would promote osteoblasts differentiation, while at the same time inhibit apoptosis. RGDS and RGES (control) peptides were covalently linked to Ti discs using an APTS linker. While the grafting of both RGDS and RGES significantly increased Ti surface roughness, contact angle analysis showed that APTS significantly increased the surface hydrophobicity; when the peptides were tethered to Ti, this was reduced. To evaluate attachment, MC3T3-E1 osteoblast cells were grown on these discs. Significantly more cells attached to the Ti-grafted RGDS then the Ti-grafted RGES control. Furthermore, expression of the osteoblasts phenotype was significantly enhanced on the Ti-grafted RGDS surface. When cells attached to the Ti-grafted RGDS were challenged with staurosporine, an apoptogen, there was significant inhibition of apoptosis; in contrast, osteoblasts adherent to the Ti-grafted RGES were killed. It is concluded that RGD-containing peptides covalently bonded to Ti promotes osteoblasts attachment and survival with minimal changes to the surface of the alloy. Therefore, such modifications to Ti would have the potential to promote osseointegration in vivo.

Nucleation and growth of calcium phosphate on amine-, carboxyl- and hydroxyl-silane self-assembled monolayers.

Upon implantation, calcium phosphate (Ca-P) surfaces form on materials that are bone bioactive. In this study, the evolving surface characteristics associated with calcium phosphate precipitation are modeled using self-assembled monolayers (SAMs), in a one-step nucleation process. SAMs were used to create amine (-NH2), carboxyl (-COOH) and hydroxyl (-OH) functionalized surfaces by grafting 3-aminopropyltriethoxysilane, 3-triethoxysilylpropyl succinic anhydride and glycidoxypropyl tri-methoxysilane, respectively, onto oxidized silicon wafers. The SAM surfaces were characterized using ellipsometry to establish the presence of grafted molecules. On the surfaces incubated in simulated physiological fluids for 7 days, the thickness of Ca-P layer grew slowly over the first few hours, increasing strongly between 1 and 5 days and then slowed down again. FTIR showed the dependence of calcium phosphate morphology on the type of surface groups, with stronger P-O bands seen on the OH-terminated surface. SEM analysis showed dispersed Ca-P precipitates on the -COOH and -OH terminated surfaces after 1 day immersion. After 7 days, all SAM surfaces were covered with uniformly dispersed and denser Ca-P precipitates. The underlying Ca-P layer showed cracks on the -NH2-terminated surface. Rutherford backscattering spectrometry (RBS) data analysis confirmed that Ca/P ratio is in excellent agreement with the theoretical value of 1.67 for hydroxyapatite. X-ray diffraction (XRD) analysis also showed evidence of apatite formation on all the surfaces, with stronger evidence on the -OH-terminated surface. Highly porous Ca-P precipitates were observed on the SAM surfaces portrayed by the AFM scans with nanoscale RMS roughness. Thus, using highly controlled surface chemistry, under physiological conditions, in vitro, this study demonstrates that a hydroxylated surface enhances Ca-P nucleation and growth relative to other surfaces, thereby supporting the concept of its beneficial effect on bone tissue formation and growth.

Osteogenic effects of bioactive glass on bone marrow stromal cells.

Bioactive glass (BG) is an effective synthetic bone graft material. BG granules of narrow size range (300-355 mum) have the ability to form new bone tissue inside excavations produced by in vivo resorption. Previously, we demonstrated that BG stimulates the differentiation of cultured osteoblast precursors if the glass surface was biomimetically modified by the formation of bone-like apatite and adsorption of serum proteins. We now report that modified BG can also increase the rate at which multipotential rat bone marrow stromal cells (rMSC) will undergo osteogenesis. BG promoted rMSC osteogenesis both when cells were plated in contact with BG and when cells were not directly in contact with the BG. Alkaline phosphatase activity, a marker of bone cell differentiation, was used as an indicator for osteogenesis. Alkaline phosphatase activity of rMSCs exposed to osteoinducers such as ascorbate, dexamethasone, and BMP-2 was enhanced in the presence of BG. The stimulatory effect of BG was more pronounced in rMSC cultures with low basal alkaline phosphatase activity than in those with higher activity. The enhanced differentiation of rMSCs was associated with both a change in rMSC morphology and altered chemical composition of the cell culture media. rMSCs cultured on BG in the presence of BMP or dexamethasone exhibited a more rounded osteoblast-like appearance as compared with cells grown on tissue culture plastic. In the presence of BG, elevated levels of calcium and silicon in the culture medium were observed throughout the 7-day culture period, suggesting a continuous dissolution of surface-modified BG and resulting release of BG dissolution products. The data suggest that both surface- and solution-mediated events play a role in the osteogenic effect of BG.

Synthesis and evaluation of novel bioactive composite starch/bioactive glass microparticles.

The aim of the development of composite materials is to combine the most desired properties of two or more materials. In this work, the biodegradable character, good controlled-release properties, and natural origin of starch-based biomaterials are combined with the bioactive and bone-bonding properties of bioactive glass (BG). Novel, bioactive composite starch-BG microparticles were synthesized starting from a blend of starch and polylactic acid (50%/50% wt) with BG 45S5 powder using a simple emulsion method. Morphological and chemical characterization showed that these particles exhibited a spherical morphology with sizes up to 350 microm and that BG 45S5 was incorporated successfully into the composite particles. Upon immersion in a solution simulating body fluids, for periods up to 3 weeks, their bioactive nature was confirmed, as a calcium-phosphate layer resembling biological apatite was formed onto their surface. The short-term cytotoxicity of these materials was also tested by placing 24-h leachables of the materials extracted in culture medium in contact with a fibroblastic cell line (L929) up to 72 h. At this time period, two biochemical tests--MTT and total protein quantification--were performed. The results showed that these materials are not cytotoxic. These results constitute the basis of future encapsulation studies using bone-acting therapeutic agents such as bone morphogenetic proteins or other bone-relevant factors. The particles developed here may be very useful for applications in which controlled release, degradability, and bone-bonding ability are the main requirements.

Model surfaces engineered with nanoscale roughness and RGD tripeptides promote osteoblast activity.

Cell adhesion to biomaterials is a prerequisite for tissue integration with the implant surface. Herein, we show that we can generate a model silica surface that contains a minimal-length arginine-glycine-aspartic acid (RGD) peptide that maintains its biological activity. In the first part of this study, attachment of MC3T3-E1 osteoblast-like cells was investigated on silicon oxide, amine terminated substrates [i.e., 3-aminopropyl triethoxysilane (APTS)], grafted RGD, and physisorbed RGD control. The APTS layer exhibited nanoscale roughness and presented amine functional groups for grafting a minimal RGD tripeptide devoid of any flanking groups or spacers. Contact angle measurements indicated that the hydrophobicity of the APTS surface was significantly lower than that of the surface with grafted RGD (RGD-APTS). Atomic force microscopy showed that surfaces covered with RGD-APTS were smoother (Ra = 0.71 nm) than those covered with APTS alone (Ra = 1.59 nm). Focusing mainly on cell morphology, experiments showed that the RGD-APTS hybrid provided an optimum surface for cell adhesion, spreading, and cytoskeletal organization. Discrete focal adhesion plaques were also observed consistent with successful cell signaling events. In a second set of experiments, smooth, monolayers of APTS (Ra = 0.1 nm) were used to prepare arginine-glycine-aspartic acid-serine (RGDS)-APTS and arginine-glycine-glutamic acid-serine (RGES)-APTS (control) substrates. Focusing mainly on cell function, integrin and gene expression were all enhanced for rate osteosarcoma cells on surfaces containing grafted RGDS. Both sets of studies demonstrated that grafted molecules of RGD(S) enhance both osteoblast-like cell adhesion and function.

In vivo evaluation of a bioactive scaffold for bone tissue engineering.

Revision cases of total hip implants are complicated by the significant amount of bone loss. New materials and/or approaches are needed to provide stability to the site, stimulate bone formation, and ultimately lead to fully functional bone tissue. Porous bioactive glasses (prepared from 45S5 granules, 45% SiO2, 24.5% Na2O, 24.5% CaO, and 6% P2O5) have been developed as scaffolds for bone tissue engineering and have been studied in vitro. In this study, we investigated the incorporation of tissue-engineered constructs utilizing these scaffolds in large, cortical bone defects in the rat simulating revision conditions. With implantation times of 2, 4, and 12 weeks the results were compared to those using the bioactive ceramic scaffold alone. Two tissue-engineered constructs were studied: osteoprogenitor cells that were either seeded onto the scaffold prior to implantation ("primary") or those that were culture expanded to form bonelike tissue on the scaffold prior to implantation ("hybrid"). Defects treated with the hybrid had the greatest amount of bone in the available pore space of the defect over all other groups at 2 weeks (p < 0.05). For both the primary and hybrid groups, woven and lamellar bone was present along the interface of the scaffold and the host cortex and within the porous space of the scaffold at 2 weeks. By 4 weeks, very uniform, lamellar bone was present throughout the scaffold for both tissue-engineered groups. The amount of bone significantly increased over time for all groups while the bioactive ceramic gradually resorbed by 40% at 12 weeks (p < 0.05). Structural properties of the treated long bones improved over time. Long bones treated with the hybrid had an early return in torsional stiffness by 2 weeks. Both tissue-engineered constructs achieved normal torsional strength and stiffness by 4 weeks as compared to the scaffold alone, which achieved this by 12 weeks. Porous, surface modified bioactive ceramic is a promising scaffold material for tissue-engineered bone repair.

Microchemical transformation of bioactive glass particles of narrow size range, a 0-24 months study.

Previous studies demonstrated the capacity of bioactive glass particles of narrow size range (300-355 microm, Biogran) to stimulate bone tissue formation without contact with pre-existing bone tissue. Chemical interactions between the bioactive glass and the surrounding tissue fluids caused the glass transformation. This study quantifies the time-dependent transformation process. Particles were implanted in the jaws of beagle dogs and resected after 1, 2, 3, 6, 12 and 24 months. Microchemical analysis was performed using a scanning electron microscope equipped with an energy dispersive X-ray analysis system. After one month, Na-ions were leached out and the particles transformed into two layers. In the center, a Si-rich gel was found on the outer surface, a Ca- and P-rich shell. After two months, the concentration levels of the outer Ca- and P-rich shell remained. In the center the Si-concentration decreased and the Ca and P concentration increased. After three months, Si disappeared completely from the center of the particle, while the Ca and P concentration increased. At one and two years, the Ca and P concentrations in the transformed particles equalled those of bone tissue, turning the transformed particle into a chemical equivalent of the bone mineral phase.

Silica sol-gel for the controlled release of antibiotics. II. The effect of synthesis parameters on the in vitro release kinetics of vanomycin.

Room temperature-processed silica sol-gel (xerogel) was investigated as a novel controlled release carrier of vancomycin for the treatment of osteomyelitis. Vancomycin-loaded xerogels were fabricated with varying water/alkoxysilane molar ratios and vancomycin concentrations. The goal of this study was to determine the effect of varying the aforementioned synthesis parameters on the daily in vitro release kinetics of vancomycin from the xerogel disks. A controlled, load-dependent, long-term release of vancomycin was observed for all of the molar ratios that were used in the study (4, 6, and 10). Variations in the water/alkoxysilane molar ratio affected the release process extensively. A cumulative release of about 90% of the original amount of vancomycin was found for molar ratios 6 and 10 by 21 and 14 days, respectively. Only about 30% was released from xerogels with a molar ratio of 4 after 21 days of immersion. A first-order release stage was followed by a steady release stage for xerogels with molar ratios of 6 and 10, whereas zero-order release was observed for xerogels with a molar ratio of 4. The findings of this study indicate that the release kinetics of vancomycin from xerogel can be tailored by varying the xerogel synthesis parameters.

Surface transformation of bioactive glass in bioreactors simulating microgravity conditions. Part I: experimental study.

Surface modified bioactive glass with surface properties akin to those of the bone mineral phase is an attractive candidate for use as a microcarrier material for 3-D growth of bone-like tissue in rotating wall vessel bioreactors (RWVs). The critical surface properties of this material are the result of reaction in solution. Because an RWV environment is completely different from conditions previously employed for bioactive glass testing, a detailed study of the surface reactions is warranted. Under properly chosen conditions, RWVs can also provide a simulated microgravity environment for the bioactive glass (BG) particles. In this sense, this study is also a report on the behavior of a bioactive material under microgravity conditions simulated on earth. A high aspect ratio vessel (HARV) and carefully selected experimental conditions enabled the simulation of microgravity in our laboratory. A complimentary numerical study was simultaneously conducted to ascertain the appropriateness of the experimental parameters (particle size, particle density, medium density, medium viscosity, and rotational speed) that ensure simulated microgravity conditions for the glass particles in the HARV. Physiological solutions (pH 7.4) with and without electrolytes, and also with serum proteins, were used to study the change in surface character resulting from simulated microgravity. Control tests at normal gravity, both static and dynamic, were also conducted. Solution and surface analyses revealed major effects of simulated microgravity. The rates of leaching of constituent ions (Si-, Ca-, and P-ions) were greatly increased in all solutions tested. The enhanced dissolution was followed by the enhanced formation of bone-like minerals at the BG surface. This enhancement is expected to affect adsorption of serum proteins and attachment molecules, which, in turn, may favorably affect bone cell adhesion and function. The findings of the study are important for the use of bioactive materials as microcarriers to generate and analyze 3-D bone-like tissue structures in bioreactors under microgravity conditions or otherwise.

Surface transformation of bioactive glass in bioreactors simulating microgravity conditions. Part II: numerical simulations.

The effects of simulated microgravity on the surface modification of bioactive glass (BG) in solution were studied using a numerical method. Models were developed for estimating the mass transfers of different chemical species from the surface of bioactive glass particles (microcarriers) suspended in the rotating liquid medium of a NASA-designed high aspect ratio vessel (HARV) bioreactor and on the bottom surface of a static vial. The concentration profiles resulting from chemical reactions and ionic transports were ascertained. Numerical results for the transport under simulated microgravity in the HARV and at normal gravity in the static vial were compared. These results were also compared with those of experiments to verify the enhancement of the reaction kinetics under simulated microgravity conditions. The experimental and numerical studies confirm that simulated microgravity conditions lead to the quick achievement of bioactive glass surface modification.

Silica sol-gel for the controlled release of antibiotics. I. Synthesis, characterization, and in vitro release.

Room temperature processed silica sol-gel (xerogel) was investigated as a novel controlled release carrier of antibiotics (vancomycin). Xerogel characteristics, in vitro release properties, and bactericidal efficacy of the released antibiotic were determined. The xerogel/vancomycin composite showed a long-term sustained release (up to 6 weeks). In addition, bactericidal efficacy of released vancomycin was retained. The kinetics of release and the amount released were dose dependent. The initial, first-order release was followed by a near-zero-order release. The time to transition from the first- to zero-order release increased with vancomycin load (from 2 to 3 weeks with load increase from 2.2 to 11.1 mg/g). Regardless of the load, about 70% of the original vancomycin content was released by the transitional point, and the cumulative release after 6 weeks of immersion was about 90%. This study, combined with other reports documenting biocompatibility and controlled resorbability of the xerogel/drug composite in vivo, suggests that silica xerogel is a promising controlled release material for the treatment of bone infections.

3D bone tissue engineered with bioactive microspheres in simulated microgravity.

Three-dimensional (3D) osteoblast cell cultures were obtained in rotating-wall vessels (RWV), simulating microgravity. Three types of bioactive microcarriers, specifically modified bioactive glass particles, bioceramic hollow microspheres, and biodegradable bioactive glass-polymer composite microspheres, were developed and used with osteoblasts. The surfaces of composite microspheres fully transformed into bone apatite after 2-wk immersion in simulated physiological fluid, which demonstrated their bone-bonding ability. The motion of microcarriers in RWVs was photographically recorded and numerically analyzed. The trajectories of hollow microspheres showed that they migrated and eventually stayed around at the central region of the RWV. At their surfaces, shear stresses were low. In contrast, solid glass or polymer particles moved toward and finally bounced off the outer wall of the RWVs. Cell culture studies in the RWV using bone marrow stromal cells showed that the cells attached to and formed 3D aggregates with the hollow microspheres. Extracellular matrix and mineralization were observed in the aggregates. Cell culture studies also confirmed the ability of the composite microspheres to support 3D bone-like tissue formation. These data suggest that the new hollow bioceramic microspheres and degradable composite microspheres can be used as microcarriers for 3D bone tissue engineering in microgravity. They also have potential applications as drug delivery systems.

45S5 bioactive glass surface charge variations and the formation of a surface calcium phosphate layer in a solution containing fibronectin.

This study investigated the effect of fibronectin adsorption on surface charge variations and calcium phosphate (Ca-P) layer formation kinetics on the surface of 45S5 bioactive glass (BG). We hypothesize that the adsorption of fibronectin on BG changes the surface charge and alters the kinetics of Ca-P layer formation on the glass surface. The charge at a material's surface modulates surface chemistry, protein adsorption, and interactions with bone cells. The zeta potential of BG in a solution containing human plasma fibronectin (TE-FN) was measured as a function of time by particle electrophoresis, and Ca-P layer formation was characterized using SEM, EDXA, and FTIR. Si, Ca, and P solution concentrations also were determined. It was found that the adsorption of fibronectin reduced the initial electronegativity of the BG surface and delayed the formation of both the amorphous and the crystalline Ca-P layers. The delayed formation of these surface layers may be attributed to the competitive binding of Ca2+ ions by the fibronectin molecule. In addition, the formation of an amorphous Ca-P layer correlated with the reversal from a negatively to a positively charged surface, independent of the presence of fibronectin. The addition of a single protein (in this case fibronectin) can significantly alter material surface parameters, such as charge, and subsequently affect the formation of a surface Ca-P layer. Furthermore, the formation of an amorphous Ca-P layer is an important event in the reactions leading to bioactive behavior, and proteins such as FN are actively involved in the transformation of the surface into a Ca-P layer.