In modular hip implants, micro-motion, which leads to fretting corrosion at the head/neck and neck/stem interfaces, has been identified as a major cause of early revision in hip implants, particularly those with heads larger than 32mm. It has been found that the type of fluid used to simulate the fretting corrosion of biomedical materials is crucial for the reliability of laboratory tests. Therefore, to properly understand and effectively design against fretting corrosion damage in modular hips, there is the need to replicate the human body environment as closely as possible during in-vitro testing and validation. In this work, corrosion behavior of CoCrMo in 0.14 M NaCl, phosphate buffered saline (PBS) and clinically relevant simulated body fluid (sbf) is carried out. Also, fretting corrosion studies of the CoCrMo alloy in a clinically relevant novel simulated body fluid (sbf) environment is studied. The presence of phosphate ions in PBS accounted for the higher corrosion rate when compared with 0.14 M NaCl and sbf environment. Despite the low and comparable corrosion rates in 0.14 M NaCl and sbf, the nature of the protective passive film formed in sbf shows the suitability of the novel sbf for future corrosion and fretting corrosion analysis. Finally, the influence of micro-motion at the modular head/neck and neck/stem interfaces on the concentration of metallic ions that goes into the synovial fluid and surrounding tissues is reported.

Made available in DSpace on 2014-06-12T15:50:51Z (GMT). No. of bitstreams: 2 arquivo5257_1.pdf: 9930279 bytes, checksum: 23011bff0745721e6391ddb8c270a9dc (MD5) license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) Previous issue date: 2006 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Compósitos feitos a partir do polímero policaprolactona e de carbonato de cálcio, chamados de PCLC, foram preparados por um processo de carbonatação, que consiste em incidir um fluxo constante de gás carbônico (CO2) em uma solução de metanol e hidróxido de cálcio por 6 horas e depois adicionar ao polímero diluído em diclorometano, após a secagem e evaporação dos solventes o material resultante é prensado no formato de pastilhas. Essas pastilhas foram então expostas, a uma solução simuladora de fluido corporal (SBF) sob temperatura constante de 37 0C, por períodos de 3, 6, 12 e 24 horas e por 7, 14, 21 e 28 dias, o que levou à deposição de estruturas do tipo apatita, semelhantes aos ossos humanos sobre sua superfície. Antes e após ser exposto ao SBF, o compósito PCLC foi analisado por diferentes técnicas de caracterização de materiais, tais como difratometria de raios-X, espectroscopia de infravermelho com transformada de Fourier (FTIR), calorimetria diferencial de varredura (DSC), microscopia eletrônica de varredura (MEV) e espectroscopia dispersiva de raios-X (EDX). Através dessa análise foi possível detectar a formação de apatitas na superfície do PCLC já a partir de 3 horas de exposição ao SBF, sendo a deposição de material inorgânico crescente com o tempo, ocorrendo variação de fases minerais e o aparecimento de hidroxiapatita após 21 dias, o que sugere a indicação do compósito PCLC como um promissor material para o desenvolvimento de implantes biocompatíveis a serem utilizados no corpo humano

This work deals with bioceramic materials based of hydroxyapatite, bioglass and their composites. These materials are commonly used in medicine, especially as hard tissue substituents. They can be prepared by different types of syntheses, from which the most common were picked for this work – precipitation of hydroxyapatite and sol-gel method for bioglass. Thermal analysis and X-ray diffraction were used for characterization of prepared powders. This thesis studies mostly their features within the composite materials, which were foamed for better bone stimulation. Properties and possibility in bio application of materials is firstly studied through their interaction in simulated body fluids, which mimics ionic concentration of human plasma. Experimental part covers synthesis of ceramic powders, their characterization, preparation of mixtures and scaffolds foamed through in situ foaming, their sintering at ideal temperatures, characterization of porosity and phase changes due to sintering. Basic tests of apatite formation ability were provided by incubation of prepared scaffolds in simulated body fluid for 3, 7, 14 and 21 days and their assay in scanning electron microscopy. Changes in concentration of Ca2+ a PO4 3- ions as well as in weight of the specimen were tracked within the incubation period.

Studies regarding the feasibility of ovalbumin (OVA) as a bone scaffold material have found its cost, availability, interaction with cells, and ability to degrade in the body into safe byproducts to be ideal for such an application. However, weak mechanical properties cause hesitation in the use of OVA as a scaffolding material in much stronger native tissue. To enhance the mechanical strength of the OVA scaffolds without compromising in vitro cellular performance, Ca-P crystals were grown on unmodified OVA and phosphonated OVA (p-OVA) samples via biomineralization processes using 5x-concentrated simulated body fluid (5x SBF). Electron microscopy (ESEM/EDS) data confirm the formation of Ca-P crystals on the surface of OVA and p-OVA scaffolds. Mechanically, rheology data measured a minimum of a three-fold increase in each mineralized scaffold's complex shear modulus over unmineralized counterparts. Degradation in a PBS+collagenase XI environment showed that mineralization extended total time to degradation. It was also shown that the formation of the Ca-P crystals had no negative effects on in vitro cell studies. To measure cellular response, a live/dead assay was conducted to confirm cell viability after 24 hours. In conclusion, improvements were made to mechanical strength without compromising in vitro cell-scaffold response. While it remains unknown whether the increase in strength is adequate for use as a bone scaffold, future work should focus on gathering necessary information to study OVA scaffolds in animal models for eventual consideration as a bone graft substitute material. / Master of Science

The overall goal of this study was to identify mineralized scaffolds which can serve as potential alternatives to bone graft substitutes intended for cleft palate repair. The aim of this preliminary study was to evaluate the role of fibrinogen (Fg) and nano-hydroxyapatite (nHA) in enhancing mineralization potential of polydioxanone (PDO) electrospun scaffolds. Scaffolds were fabricated by blending PDO:nHA:Fg in the following weight ratios: 100:0:0, 50:25:25, 50:50:0, 50:0:50, 0:0:100 and 0:50:50. Scaffolds were immersed in different simulated body fluids for 5 and 14 days to induce mineralization. The inclusion of fibrinogen induced sheet-like mineralization while individual fiber mineralization was noticed in its absence. Modified protocols of alizarin red staining and burn-out test were developed to quantify mineral content of scaffolds. After mineralization, 50:50:0 scaffolds were still porous and contained the most mineral. 50:25:25 scaffolds had the highest mineralization potential but lacked porosity. Therefore, it can be anticipated that these mineralized organic-inorganic electrospun scaffolds will induce bone formation.

Apatite was formed on the surface of titanium dioxide (TiO2) powders by a biomimetic process. The deposition was accomplished by immersing TiO2 powders in simulated body fluid (SBF) for 1, 3, 6, 12, and 24 h. SBF used throughout this study had calcium and phosphate ion concentrations 10 times greater than those of human blood plasma. Photocatalytic activity of the apatite-deposited TiO2 powders was investigated in terms of the decomposition of methylene blue solution under ultraviolet (UV) irradiation. It has been shown that apatite deposition enhanced the photocatalytic activity of TiO2. The best photocatalytic performance was acquired on the powders that are immersed in SBF for 3 h. The time required for the complete degradation of methylene blue decreased from 3.5 h to 2 h upon immersion of powders in SBF for 3 h. Photochemical durability of poly(methyl methacrylate) increased when it was mixed with apatite-deposited TiO2 powders.

Tissue Engineering is a scientific discipline that aims to regenerate tissues and organs that are diseased, lost or congenitally absent. It encompasses the use of suitable synthetic equivalents of native extracellular matrix that may or may not be supplemented with cells or relevant growth factors. Such scaffolds are designed to reside at the site of implantation for a variable period of time during which they induce the regeneration of native tissue. During this time, they also provide a template for new cells to attach, infiltrate, differentiate into appropriate phenotype and eventually restore function of the concerned tissue. Among the factors that affect the outcome are the composition of scaffold, methods of fabrication, bulk properties of the scaffold and topography and architecture at the cellular level. Bone is unique in the body in that it is one of the few tissues capable of complete regeneration even in adults, as seen during fracture healing. However, certain conditions (non-union of fractures, congenital and acquired bone deficiencies) exist in which the regenerative capacities of bone are exceeded and appropriate intervention becomes necessary. Current treatment options include autologous bone grafts harvested from iliac crest or de-cellularized allografts or synthetic substitutes made from metals, ceramics and polymers. However these options have serious limitations: while autografts are limited in supply, necessitate second surgery and show inadequate vascularization, allografts can transmit viral infections. Metals, ceramics and polymers are in essence structural replacements without performing any biological function. Other problems associated with these synthetic materials include adverse immune reactions, corrosion, stress-shielding and secondary fractures due to inadequate osseo-integration. Bone tissue engineering is a specialized field of research that provides an alternative strategy to repair bone defects by exploiting the advances in engineering and better understanding of bone biology. Scaffold-based tissue engineering approach is a promising field that involves implantation of a biomaterial that is specifically matched in terms of biological and material properties to the tissue it replaces. This study explores the feasibility of using electrospinning as a potential fabrication strategy for bone tissue engineering applications, more specifically intended for cleft palate repair. This model represents a congenital deformity that affects both hard and soft tissues and presents unique challenges and opportunities. Among the challenges are: the need for the implant allow growth of the most complex areas of the facial skeleton, integrate and grow with the patient through adolescence, the ability of the implant to not interfere with vital functions including breathing and feeding. Further the implant should provide a flexible matrix that can effectively support erupting teeth. In spite of these extreme demands, maxilla is a non load-bearing membranous bone, a favorable consideration from materials engineering perspective. The present study is organized into three independent sections. The first section investigates developing strategies intended to improve the material properties of electrospun bone scaffold. Bone is composed of a high volume fraction (50%) of inorganic hydroxyapatite nanocrystals that is closely associated with collagen. The dispersal of brittle mineral is critical in not only strengthening the bone in compression but also contributes to the osteoconductivity of the matrix. Since loading of mineral in a bone scaffold is a serious limitation, we attempted to achieve improved loading of bone mineral by dual mineralization approach. We first incorporated nanocrystalline hydroxyapatite (nHA) directly into the scaffold by adding it to the electrospinning polymer solution. The second step involves inducing biomimetic mineralization of electrospun scaffolds by incubating them in simulated body fluid (SBF) for 2 weeks. The hypothesis was that the nanocrystalline hydroxyapatite seeded during electrospinning would act as sites for nucleation and further crystal growth when incubated in solution supersaturated with respect to calcium and phosphate ions. We tested this approach in two synthetic, biocompatible polymers-polydioxanone and poly (lactide: glycolide) and four formulations of SBF with differential loading of nHA (0-50% by wt. of polymer). A modified Alizarin Red S (ARS) staining that specifically binds to calcium was developed that allowed us to quantify the mineral content of 3D scaffold with great accuracy. Results indicated a unique combination of factors: PDO scaffolds containing 50% nHA incubated in 1x revised-SBF incubated under static conditions gave maximum mineralization over a period of two weeks. We then sought to exploit these findings to engineer a stiffer scaffold by stacking multiple layers together and cold welding them under high pressure. Electrospun scaffolds (1, 2 or 4 layered stacks) were either compressed before or after mineralizing treatment with SBF. After two weeks, scaffolds were analyzed for total mineral content and stiffness by uniaxial tensile testing. Results indicated while compression of multiple layers significantly increases the stiffness of scaffolds, it also had lower levels of mineralization partly due to increased density of fibers and loss of surface area due to fiber welding. However this can be offset to a reasonable degree by increasing the number of stacks and hence this strategy can be successfully adopted to improve the mechanical properties of electrospun scaffolds. The second section introduces a novel infrared imaging technique to quantify and characterize the biological activity of biomaterials, based on cell adhesion. Cells attach to the surface by the formation of focal contacts where multiple proteins including vinculin and talin assemble to signal critical processes like cell survival, migration, proliferation and differentiation. After allowing MG-63 osteoblasts to adhere to 2D biomaterial surface coated with extracellular matrix proteins (collagen, gelatin, fibronectin) cells were fixed and probed with antibodies for vinculin and talin. Secondary antibodies, tagged with infrared-sensitive fluorescent dyes, were used to quantify the molecules of interest. In addition, the kinetics of focal contact formation in these different substrates was followed. Successful quantification of focal contacts were made and further research revealed phosphorylation of vinculin at pY-822 as one potential mechanism for recruitment of vinculin to focal contacts. Hence it could represent a subset of vinculin and might serve as a specific molecular marker for focal contacts. As an extension, we evaluated the possibility of using such an assay to quantify 3D electrospun tissue engineering scaffolds. We fabricated scaffolds of graded biological activity by electrospinning blends of polydioxanone and collagen in different ratios. Vinculin and talin expressed by MG-63 cultured on these scaffolds for 24 hours were quantified in a similar manner. Results indicate that while talin does not show a significant difference in expression among different scaffolds, vinculin showed a positive correlation with increasing biological activity of scaffolds. In conclusion, we have identified vinculin as a reliable marker of focal contacts in 3D scaffolds while phosphovinculin (pY-822) was more specific to focal contacts in coated 2D substrates. In both instances, infrared imaging proved to be reliable in study of focal contacts. The third section aims to make the bone scaffolds osteoinductive- a property of a material to induce new bone formation even when implanted in subcutaneous and intramuscular heterotopic sites. Bone morphogenetic proteins (BMP) are potent cytokines that can induce migration, proliferation and differentiation of stem cells along osteoblastic lineage. The therapeutic efficacy of BMPs in the treatment of severe bone defects has been identified and is currently FDA approved for specific orthopedic applications. BMPs are clinically administered in a buffer form that not only makes the treatment expensive but less effective. Suitable delivery systems for BMP delivery have been an intense area of investigation. We rationalized electrospinning as a strategy to incorporate BMP within the scaffold and that would enable controlled release when implanted. One of the drawbacks of using electrospinning to deliver bioactive molecules is the potential denaturing effect and eventual loss of activity of BMPs. The final section of this dissertation tries to develop sensitive and relevant assays that could answer intriguing questions about solvent-protein interaction. We chose to use the BMP-2/7 heterodimer as the osteoinductive molecule of choice because of its superior potency compared to homodimer counterparts. We characterized the detection and quantification of BMP-2/7 using a slot blot technique. Further, we used a novel cell line (C2C12 BRA) to test the retention of activity of BMP-2/7 that has been exposed to organic solvents. Results indicate significant loss of activity when BMPs are exposed to organic solvents but complete recovery was possible by diluting the solvent with an aqueous buffer.

Pour combiner les bonnes propriétés mécaniques des matériaux métalliques et obtenir des surfaces bioactives, des revêtements de phosphate de calcium (Ca-P) ont été développés. Le but est d’améliorer la biocompatibilité et la bioactivité du système et créer une barrière contre l’éventuelle libération d’ions toxiques du substrat métallique. Notre démarche a été de développer une nouvelle voie de synthèse. Après un polissage mécanique et décapage chimique du substrat de Titane ou Ti–6Al–4V une couche intermédiaire en titanate de sodium a été obtenue par un prétraitement alcalin sur les substrats. Elle a été ensuite soumise à un traitement thermique afin de créer une couche alvéolaire nanométrique. Celle-ci facilite la croissance une couche de phosphate de calcium par voie autocatalytique d'une manière similaire au procédé de formation de l'os naturel. La caractérisation et l’étude des revêtements des Ca-P obtenus par les trois bains (acides, alcalins et oxydant) sur les substrats métalliques ont été réalisés. Les dépôts ont été étudiés d’un point de vue structural et morphologique. La stabilité des couches de Ca-P a été mesurée dans le fluide corporel (SBF) pour des périodes différentes utilisant des analyses biochimiques. En conclusion, cette méthode est une solution peu coûteuse, fiable, et utilisable à l'échelle industrielle. La couche de Ca-P aussi bien que le titanate de sodium obtenus peuvent permettre d’être imprégnés par des agents actifs comme un ou plusieurs agents antibactériens. / To combine the good mechanical properties of metallic substrate with the bioactivity of some ceramics, to obtain bioactive surface, a calcium phosphate (Ca-P) coating has been developed. The goal is to improve the biocompatibility and bioactivity of metallic implant and create a barrier against the possible release of toxic ions from the metallic substrate. Our approach has been to develop a new synthetic route. After mechanical polishing and chemical etching of the titanium or Ti-6Al-4V substrates, an interlayer of sodium titanate was obtained by an alkaline pretreatment of the substrates. Then, it was subjected to a heat treatment to create a nanometeric layer. The latter facilitates the growth of a layer of Ca-P by means of an electroless manner similar to the forming process of the natural bone. Characterization and study of Ca-P coatings obtained by the three baths (acids, alkalis and oxidizing) on metallic substrates have been made. The deposits were studied from a structural and morphological point of view. The stability of Ca-P layers was measured in simulated body fluid (SBF) for different periods using biochemical analyzes. In conclusion, this method is a cheap and reliable solution that can be used on an industrial scale. The Ca-P as well as the sodium titanate layers can afford to be impregnated with active agents such as one or more antibacterial agents.

This thesis deals with the characterization of electrochemical corrosion properties of magnesium alloys as promising materials for biomedical applications. The wrought alloys AZ31 and AZ61 were used and exposed to corrosive environments of Hanks solutions (SBF) to simulate environmental conditions in living organisms. For the evaluation of the surfaces was used scanning electron microscopy (SEM) with elemental analysis measured by energy-dispersive spectroscopy (EDS). Short-term (5 min) and long-term (72 h) corrosion tests were conducted in order to optimize the measurement methodology and obtain corrosion parameters - especially corrosion potential (Ekor), corrosion current density (ikor) and polarisation resistance (RP). To evaluation of the short-term tests were by potentiodynamic tests, namely the linear polarization (LP) test. Long-term tests were measured by electrochemical impedance spectroscopy (EIS). Effects of the composition of the alloys (AZ31 and AZ61), surface treatment (grinding and polishing) and the composition of the solution (SBF without Ca, Mg, and with Ca, Mg) were compared. Complex corrosion behaviour in time was characterized and corrosion mechanisms were discussed.

Se sintetizaron nanocompuestos híbridos en bloque de poli(etil metacrilato-co-hidroxietil acrilato) 70/30 wt%/sílice, P(EMA-co-HEA)/SiO2, con distintas proporciones de sílice hasta el 30 wt%. El procedimiento de síntesis consistió en la copolimerización de los monómeros orgánicos durante la polimerización sol-gel simultánea de tetraetoxisilano, TEOS como precursor de sílice. El TEOS se hidroliza eficientemente y condensa dando lugar a sílice, y presenta una distribución homogénea en forma de agregados inconexos de nanopartículas de sílice elementales en los híbridos con bajos contenidos de sílice (<10 wt%) o redes continuas interpenetradas con la red orgánica tras la coalescencia de los agregados de sílice (>10 wt%). La red polimérica orgánica se forma en los poros producidos en el interior de las nanopartículas elementales de sílice, y también en los poros formados entre los agregados de nanopartículas. Los nanohíbridos con contenidos de sílice intermedios (10-20 wt%) exhibieron las propiedades más equilibradas e interesantes: i) refuerzo mecánico de la matriz orgánica conseguida gracias a redes de sílice continuas e interpenetradas, ii) buena capacidad de hinchado debida a la expansión de la red orgánica no impedida todavía por un esqueleto de sílice rígido, y a un número alto de grupos silanol terminales hidrófilos (concentraciones inorgánicas en los alrededores de la coalescencia), y iii) mayor reactividad superficial debido a un contenido relativo bastante elevado de grupos polares silanol terminales disponibles en las superficies. La 'bioactividad' o capacidad de los materiales en bloque de formar hidroxiapatita (HAp) sobre sus superficies fue estudiada in vitro sumergiéndolos en fluido biológico simulado (simulated body fluid, SBF). La formación de la capa de HAp viene controlada por el mecanismo y el tiempo de inducción a la nucleación de la misma, que dependen a su vez de la estructura de la sílice. / Vallés Lluch, A. (2008). P(EMA-co-HEA)/SiO2 hybrid nanocomposites for guided dentin tissue regeneration: structure, characterization and bioactivity [Tesis doctoral no publicada]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/3795 / Palancia