[pt] A hidroxiapatita (HA) é um constituinte natural dos ossos e rochas ígneas e tem grande importância na reserva de cálcio e fósforo dos vertebrados. Possui propriedades de biocompatibilidade e bioatividade, permitindo que haja substituições de seus átomos por outros. Neste trabalho são estudadas as substituições de Cd e Sr nos dois sítios cristalinos, 1 e 2, de Ca. São apresentados cálculos de estrutura eletrônica (ordem de ligação e carga atômica) em aglomerados de HA, Cd/HA e Sr/HA, utilizando o método de Huckel estendido (eHT), que se baseia na aproximação de ligações fortes (tight binding) em base não ortogonal. Os parâmetros empíricos utilizados foram ajustados de modo a reproduzir dados experimentais e cálculos ab initio nos óxidos CaO, CdO e SrO. Os resultados para os sistemas HA mostram que, em ambas as substituições, o sítio 2 é o preferencial. São feitas comparações com resultados encontrados na literatura, tanto experimentais quanto calculados através da Teoria do Funcional Densidade(DFT). O presente estudo corrobora trabalhos anteriores mostrando que o método semi-empírico eHT pode fornecer uma descrição qualitativa adequada das propriedades eletrônicas de sistemas envolvendo HA pura ou com substituições. / [en] Hydroxyapatite (HA) is a natural constituent of bones and igneous rocks. It plays important role in the storage of calcium and phosphorus of vertebrates. HA presents biocompatibility and bioactivity properties, allowing substitutions of its atoms for others. In this work, the substitutions of Cd and Sr in two crystal sites, 1 and 2, of Ca, are studied. Electronic structure calculations (bond order and atomic charge) are performed on pure and Cd/HA and Sr/HA clusters, by using the extended Hückel Theory (eHT), based on non orthogonal tight binding approximations. For a better description of the HA systems, it is used empirical parameters adjusted in order to reproduce known band structure and experimental data on CaO, CdO and SrO oxides. It is shown that site 2 is the preferred substitution site for both, Cd and Sr. This result is discussed in connection with previous experimental and DFT based calculations on the two substituted HA. The present study corroborates previous work which suggested that eHT may provide a reasonable qualitative description of basic electronic properties of both pure and substituted hydroxyapatite materials.

Cochlear implants (CIs) help to restore basic auditory function in patients who are deaf or have profound hearing loss. However, CI patients suffer from limited voice and tonal perception due to spatial separation between the stimulating CI electrode and the receptor spiral ganglion neurons (SGNs). Directed regeneration of proximate SGN axons may improve tonal performance and implant fidelity by decreasing the spatial separation between the CI electrode and the neural receptor. Additionally, fibrous scar tissue formation on the surface of implanted electrodes further decreases tonal perception through current attenuation and spreading resulting in late-term hearing loss. Thus, designing surfaces that induce favorable responses from neural tissues will be necessary in overcoming signal resolution barriers. In this work, the inherent spatial and temporal control of photopolymerization was used to functionalize surfaces with topographical and biochemical micropatterns that control the outgrowth of neural and other cell types. First, laminin, a cell adhesion protein was patterned using a photodeactivation process onto methacrylate polymer surfaces and was shown to direct the growth of spiral ganglion neurons (SGN), the primary auditory neural receptors. These protein patterns could even overcome low amplitude/high periodicity competing topographical cues. Additionally, glass substrates were patterned with linear zwitterionic polymers and fibroblasts, astrocytes, and Schwann cells all showed dramatically decreased cell adhesion on 100 µm precocity patterns. Further, SGN neurites showed excellent alignment to these same patterns. Next, poly(dimethyl siloxane) (PDMS) was coated with a crosslinked zwitterionic thin film using a single step photografting/photopolymerization process to covalently bind the hydrogel to PDMS. These coated surfaces showed dramatically lower levels of protein, cell, and bacterial adhesion. Finally, zwitterionic hydrogels were strengthened by changing the concentration of poly(ethylene glycol) diacrylate (PEGDA) and 2-hydroxyethyl methacrylate (HEMA) in the formulation. The direct relationship between changing zwitterionic hydrogel formulation to strengthen the hydrogel and the anti-fouling properties were established. The fundamental understanding and design of cochlear implant materials described herein serves as a foundation for the development of next generation neural prosthetics.

The overall thrust of this dissertation is to gain a fundamental understanding of the synergistic effects between surface topography and chemical functionality of poorly adhesive materials on enhancing the adhesion of mouse embryonic fibroblasts. Cellular response to surface topography and chemical functionality have been extensively studied on their own providing valuable information that helps in the design of new and improved biomaterials for tissue engineering applications. However, there is a lack of understanding of the synergistic effect of microscale and nanoscale topography with chemical functionality and the relative impact and contribution of each in modulating cellular behavior. By understanding the relationship between these cues, in particular using materials that are poorly adhesive, this study will provide new clues as to how cells adapt to their environment and also suggest new dimensions of biomaterial design for fine-tuning cellular control. A microstructure that combined non adhesive materials with defined surface topography and surface chemistry is presented, to assess and correlate the enhancement of mouse embryonic fibroblasts cell adhesion and spreading. Poly (N-isopropylacrylamide) or PNIPAAm electrospun fibers were overlaid on PNIPAAm thin films (100 nm) at various time points to investigate the role of topography on such coatings by keeping the chemical functionality the same. After doing this, several topographical patterns were developed, spanning from sparse to dense fiber mats, and cell adhesion strongly depended on the relative available areas for attachment on either the fibers or the supporting surface. To gain a better understanding of this finding, two surface chemistries, non-adhesive (self-assembled monolayer of polyethylene glycol (PEGSAM) alkanethiol on gold) or an adhesive coating (3-aminopropyltriethoxysilane (APTES) on glass) with well characterized adhesive properties were included in this study to assess the effect of topographical cues provided by the PNIPAAm electrospun fibers on cellular responses. With the deposition of the PNIPAAm fibers onto a PEGSAM surface, cell adhesion increased to almost 100%, and unlike the PNIPAAm surface, cell spreading was significantly enhanced. With the deposition of PNIPAAm fibers onto APTES, both cell adhesion and spreading were unaffected up to 60% fiber coverage. For both surfaces, PNIPAAm fiber densities above 60% coverage lead to adhesion and spreading independent of the underlying surface. These findings indicate the presence of a sparse topographical feature can stimulate cell adhesion on a typically non-adhesive material, and that a chemical dissimilarity between the topographic features and the background enhances this effect through greater cell-surface interaction. In addition to the aforementioned studies, cell response was also assessed on PNIPAAm thin films coatings with thicknesses ranging from 100 nm to 7 nm. Cell adhesion and spreading was enhanced as the thickness of the thin film decreased. This change was more noticeable below 30 nm, wherein 7 nm shows the highest cell adhesion and spreading enhancement. The results reported are preliminary results and further experiments will be conducted, to support the data. It is believed that cellular response was enhanced due to a change in surface topography at the nanoscale level.

In vivo, les cellules souches résident au sein de niches, des microenvironnements extrêmement spécialisés qui leur permettent de réguler leur prolifération ainsi que leur différentiation en cellules matures et fonctionnelles. Ces microenvironnements sont caractérisés par des interactions cellules-cellules, la présence de facteurs de croissance et de cytokines, et une matrice extracellulaire, qui fournissent des signaux qui permettent de contrôler le comportement des cellules souches.Bone morphogenetic protein 2 (BMP-2) est un facteur de croissance ostéoinductif qui est impliqué dans toutes les étapes de la différentiation ostéoblastique, ainsi que dans la transdifférentiation de myoblastes vers une lignée ostéogénique. In vivo, ce facteur existe à la fois en solution et lié à la matrice extracellulaire. Bien que ces deux modes de présentation influencent les comportements cellulaires de façon distincte, leur rôle dans le microenvironnement de la niche et leur pertinence fonctionnelle dans la genèse de la réponse biologique n’a presque pas été étudié à l’échelle cellulaire. Ici nous avons utilisé l’affinité naturelle de la BMP-2 pour la fibronectine afin de créer des micromotifs de BMP-2 de taille cellulaire sur des biomatériaux mous.Cette technique nous a permis de contrôler simultanément la présentation spatiale de la BMP-2 liée à la fibronectine, ainsi que l’étalement cellulaire. Ces micromotifs ont induit une organisation d’actine et d’adhésions focales autour du noyau, et a déclenché la phosphorylation et la translocation nucléaire de SMAD1/5/8 dans des myoblastes murins C2C12 et des cellules souches mésenchymateuses. Ceci est un indicateur précoce de leur transdifférentiation ostéoblastique. Nous avons trouvé que l’étalement cellulaire lui-même facilitait la phosphorylation de SMAD1/5/8 dépendante de la BMP-2, et avons démontré que la signalisation SMAD médiée par la fibronectine et la BMP-2 dépendait de LIM kinase 2 et de ROCK, plutôt que d’une activation de la myosine II.Nous avons également pu utiliser cet outil pour étudier les cinétiques d’adhésion et d’étalement, les changements d’organisation du cytosquelette dépendants du mode de présentation de la BMP-2, et la signalisation entre la matrice et la cellule, médiée par les intégrines. Nous avons obtenu des résultats préliminaires suggérant un effet du mode de présentation de la BMP-2 sur les forces cellulaires, suggérant que le mode de présentation des facteurs de croissance pourrait être pertinent à d’autres mécanismes cellulaires tels que la mécanotransduction.Dans l’ensemble, nous résultats montrent que les patterns de BMP-2 liée à la fibronectine sont un outil utile à l’étude de mécanismes cellulaires. De façon plus large, notre approche pourrait être adaptée à d’autres combinaisons de protéines de la matrice et de facteurs de croissance, ouvrant ainsi une avenue fascinante pour recréer in vitro des niches spécifiques aux tissus. / In vivo, stem cells reside within niches, highly specialized microenvironments which allow them to regulate their self-renewal and differentiation into mature and functional cells. These microenvironments are characterized by cell-cell interactions, the presence of growth factors and cytokines, and an extracellular matrix, all of which provide cues that control stem cell behavior.Bone morphogenetic protein 2 (BMP-2) is an osteoinductive growth factor which is involved in all stages of osteoblastic differentiation, as well as the transdifferentiation of myoblasts toward an osteogenic lineage. In vivo, it exists both in solution and bound to the ECM. While these two modes of presentation are known to influence cell behavior distinctly, their role in the niche microenvironment and their functional relevance in the genesis of a biological response has sparsely been investigated at a cellular level. Here we used the natural affinity of BMP-2 for fibronectin (FN) to engineer cell-sized micropatterns of BMP-2 on soft biomaterials.This technique allowed the simultaneous control of the spatial presentation of fibronectin-bound BMP-2 and cell spreading. These micropatterns induced a specific actin and adhesion organization around the nucleus, and triggered the phosphorylation and nuclear translocation of SMAD1/5/8 in C2C12 myoblasts and mesenchymal stem cells, an early indicator of their osteoblastic transdifferentiation. We found that cell spreading itself potentiated a BMP-2-dependent phosphorylation of SMAD1/5/8, and demonstrated that FN/BMP-2-mediated early SMAD signaling depended on LIM kinase 2 and ROCK, rather than myosin II activation.We were also able to use this tool to investigate adhesion and spreading kinetics, changes in cytoskeletal organization depending on BMP-2 presentation mode, and reciprocal integrin-mediated signaling between the ECM and the cell. We were able to show preliminary results suggesting an effect of BMP-2 presentation mode on cellular forces, suggesting that growth factor presentation may be relevant to other cellular mechanisms like mechanotransduction.Altogether, our results show that FN/BMP-2 micropatterns are a useful tool to study the mechanisms underlying BMP-2-mediated mechanotransduction. More broadly, our approach could be adapted to other combinations of ECM proteins and growth factors, opening an exciting avenue to recreate tissue-specific niches in vitro.

<p>Today there are a vast number of medical devices in temporary or permanent contact with human tissues. Blood-biomaterial contact is known to trigger the complement system and results in generation of fluid phase anaphylatoxins C3a and C5a, and surface-bound C3b and iC3b. All these products together are able to attract and activate leukocytes and trigger release of inflammatory mediators leading to a systemic inflammation indirectly causing hemostatic problems and even organ failure. The aim of this study was to identify how complement is triggered on a biomaterial surface and to find ways to regulate this activation.</p><p>The finding that complement activation on biomaterials can be divided into initiation and amplification will facilitate regulation of complement activation biomaterial surfaces. This concept is also compatible with the two techniques to regulate complement activation on a surface.</p>

Today there are a vast number of medical devices in temporary or permanent contact with human tissues. Blood-biomaterial contact is known to trigger the complement system and results in generation of fluid phase anaphylatoxins C3a and C5a, and surface-bound C3b and iC3b. All these products together are able to attract and activate leukocytes and trigger release of inflammatory mediators leading to a systemic inflammation indirectly causing hemostatic problems and even organ failure. The aim of this study was to identify how complement is triggered on a biomaterial surface and to find ways to regulate this activation. The finding that complement activation on biomaterials can be divided into initiation and amplification will facilitate regulation of complement activation biomaterial surfaces. This concept is also compatible with the two techniques to regulate complement activation on a surface.

<p>This thesis presents an analysis of the material properties and phase behavior of divalent ionic surfactant salts, and protein and polymer glasses. There has been extensive interest in understanding the phase behavior of divalent ionic surfactants due to the many applications of ionic surfactants in which they come into contact with divalent ions, such as detergency, oil recovery, and surfactant separation processes. One goal of determining the phase boundaries was to explore the option of incorporating a hydrophobic molecule into the solid phase through the micelle-to-crystal bilayer transition, either for drug delivery applications (with a biologically compatible surfactant) or for the purpose of studying the hydrophobic molecule itself. The liquid micellar and solid crystal phases of the alkaline earth metal dodecyl sulfates were investigated using calorimetry, visual inspection, solubilization of a fluorescent probe, and x-ray diffraction. The Krafft temperature and dissolution enthalpy were determined for each surfactant, and partial composition-temperature phase diagrams of magnesium dodecyl sulfate-water, calcium dodecyl sulfate-water, as well as sodium dodecyl sulfate with MgCl₂ and CaCl₂ are presented. As a proof of concept, fluorescence microscopy images showed that it is, in fact, possible to incorporate a small hydrophobic molecule, diphenylhexatriene, into the solid phase.</p><p>The second, and main, part of this thesis expands on work done previously in the lab by using the micropipette technique to study two-phase microsystems. These microsystems consist of a liquid droplet suspended in a second, immiscible liquid medium, and can serve as direct single-particle studies of drug delivery systems that are formed using solvent extraction (e.g., protein encapsulated in a biodegradable polymer), and as model systems with which to study the materials and principles that govern particle formation. The assumptions of the Epstein-Plesset model, which predicts the rate of droplet dissolution, are examined in the context of the micropipette technique. A modification to the model is presented that accounts for the effect a solute has on the dissolution rate. The modification is based on the assumption that the droplet interface is in local thermodynamic equilibrium, and that the water activity in a solution droplet can be used to determine its dissolution (or dehydration) rate. The model successfully predicts the dissolution rates of NaCl solutions into octanol and butyl acetate up to the point of NaCl crystallization. The dehydration of protein solutions (lysozyme or bovine serum albumin) results in glassified microbeads with less than a monolayer of water coverage per protein molecule, which can be controlled by the water activity of the surrounding organic medium. The kinetics of dehydration match the prediction of the activity-based model, and it is shown how the micropipette technique can be used to study the effect of dissolution rate on final particle morphology. By using a stable protein with a simple geometry (lyosyzme), this technique was be used to determine the distance dependence of protein-protein interactions in the range of 2-25 &Aring;, providing the first calculation of the hydration pressure decay length for globular proteins. The distance-dependence of the interaction potential at distances less than 9 &Aring; was found to have a decay length of 1.7 &Aring;, which is consistent with the known decay length of hydration pressure between other biological materials. Biodegradable polyesters, such as poly(lactide-co-glycolide) (PLGA), are some of the most common materials used for the encapsulation of therapeutics in microspheres for long-term drug release. Since they degrade by hydrolysis, release rates depend on water uptake, which can be affected by processing parameters and the material properties of the encapsulated drug. The micropipette technique allows observations not possible on any bulk preparation method. Single-particle observations of microsphere formation (organic solvent extraction into a surrounding aqueous phase) show that as solvent leaves the microsphere and the water concentration in the polymer matrix becomes supersaturated, water phase separates and inclusions initially grow quickly. Once the concentration in the polymer matrix equilibrates with the surrounding aqueous medium, the water inclusions continue to grow due to dissolved impurities, solvent, and/or water-soluble polymer fragments resulting from hydrolysis, all of which locally lower the water activity in the inclusion. Experiments are also presented in which glassified protein microbeads were suspended in PLGA solution prior to forming the single microspheres. This technique allowed the concentration of protein in a single microbead/inclusion to be determined as a function of time.</p> / Dissertation

Tissue engineering is a multidisciplinary subject aimed at producing the immunologically tolerant artificial tissues/organs to repair or replace damaged ones. In this field, tissue scaffold plays a key role to support cell growth and new tissue regeneration. For fabrication of tissue scaffolds with individual external geometry and predefined inner structure, rapid prototyping (RP) systems based on fluid dispensing techniques have proved to be very promising. The present research conducted a comprehensive study on the dispensing-based fabrication process. <p>First of all, the scaffold materials are characterized in terms of their biocompatibility and flow behaviour. The biocompatibility of biomaterials of PLLA, PCL, collagen, chitosan, and gelatine is evaluated in terms of supporting neuron cells adhesion and outgrowth. Chitosan solution (2% w/v) in acetic acid is shown to be the most promising among the examined biomaterials for the fabrication of nerve tissue scaffolds. Its non-Newtonian flow behaviour is identified by using a commercial rheometer. <p>In the fabrication process, the flow rate of biomaterials dispensed, the profile of strand cross-sections, and the scaffold porosity are very important and must be precisely controlled. A model is developed to represent the flow rate of biomaterials dispensed under the assumptions that the flow is incompressible, steady, laminar, and axisymmetric. Also, the profile and size of line strands at different layers and portions are modeled based on the Young-Laplace equation. Thus the dispensing-based fabrication process can be predicted in terms of the flow rate and the scaffold porosity. <p>The effects of operation conditions on the fabrication result are identified theoretically and experimentally. Simulation result shows that a higher driving pressure, a higher temperature, and a larger needle diameter will result in a larger size of the strand cross-sections and lower scaffold porosity. The change pattern, however, is nonlinear, which is affected by the fluid surface tension and non-Newtonian flow behaviour of scaffold biomaterials. <p>To verify the effectiveness of the developed models, experiments were carried out on a commercial dispensing system (C-720, Asymtek, USA). To avoid the possible error derived from the temperature difference between the dispensing system and the rheometer, a new method is presented to characterize the fluid properties used for model predictions. Experimental results illustrate that the developed models, combined with the new identification method, are very promising to predict the dispensing-based fabrication process.

The field of Tissue Engineering has developed in response to the shortcomings associated to the replacement of tissues lost to disease or trauma: donor tissue rejection, chronic inflammation, and donor tissue shortages. The driving force behind Tissue Engineering is to avoid these problems by creating biological substitutes capable of replacing the damaged tissue. This is done by combining scaffolds, cells and signals in order to create living, physiological, three-dimensional tissues.Scaffolds are porous biodegradable structures that are meant to be colonised by cells and degrade in time with tissue generation. Scaffold design and development is mainly an engineering challenge, and is the goal of this PhD thesis.The main aim of this thesis is to develop and characterise scaffolds for Tissue Engineering applications. Specifically, its objectives are: 1. To study, optimise and characterise two scaffold processing methods: Solvent Casting and Phase Separation. This is done by experiment design analysis. 2. To characterise the degradation, surface properties, and cellular behaviour of the scaffolds produced. The scaffolds are made of a composite of polylactic acid polymer and a calcium phosphate soluble glass. The comparison of the two processing methods reveals that in general, the solvent cast scaffolds have higher porosities and lower mechanical properties than the phase-separated ones. Two compositions containing 20 weight % and 50 weight % of glass particles were chosen for further characterisations including degradation, surface properties and cellular behaviour. The degradation of the scaffolds was studied for a period of 10 weeks. The evolution of various parameters such as: morphology, weight loss, mechanical properties, thermal transitions and porosity, was monitored. Scaffolds produced via solvent casting were found to be more severely affected by degradation than phase-separated ones. The surface properties of the scaffolds were measured by modelling the scaffold pore walls as thin composite films. The morphology, topography, surface energy and protein adsorption of the films was characterised thoroughly. Again, the processing method was critical in determining scaffold properties. Films made via phase-separation processing had markedly different properties due to extensive coating of the glass particles by the polymer. This made the surfaces rougher and more hydrophobic. When the glass particles are not completely coated with polymer, they increase the material's hydrophilic and protein adsorption properties, thus confirming the potential biological benefits of the inclusion of the calcium phosphate glass.The biological behaviour of the scaffolds was characterised by means of in vitro cell cultures with primary osteoblast stem cells and cells from a stable cell line, under static and dynamic conditions. Their morphology, proliferation and differentiation were monitored. Both types of scaffolds sustained osteblastic cell growth. The solvent cast scaffolds were easily colonised by cells which migrated throughout their structure. The cells on the phase-separated scaffolds, however, tended to form thick layers on the scaffold surface. Finally, an alternative characterisation technique was explored applying Synchrotron X-Ray Microtomography and in-situ micromechanical testing. These experiments allowed for the qualitative and quantitative analysis of the microstructure of the scaffolds both at rest and under strain. A finite element model of the solvent cast scaffolds was developed and a preliminary analysis was performed. This technique could be used to complement and overcome some of the limitations of traditional mechanical characterisation of these highly porous materials.

Un campo que despierta un interés creciente dentro del ámbito de los biomateriales para la regeneración osea es el de los cementos de fosfatos de calcio. Un cemento de fosfato de calcio consiste en una fase sólida formada por polvos de fosfatos de calcio, y una fase liquida, que al ser mezcladas fraguan a temperatura ambiente o fisiológica, dando lugar a un cuerpo sólido. Junto a la biocompatibilidad y bioactividad de las cerámicas de fosfatos de calcio, los cementos presentan una serie de ventajas especificas, derivadas de su proceso de obtención, como la moldeabilidad y la capacidad de fraguar "in situ", que pueden resolver los problemas de fijación y adaptación planteados por las cerámicas de fosfatos de calcio.En este trabajo se desarrolla y caracteriza un cemento basado en la hidrólisis del fosfato tricalcico alfa. El fraguado del cemento se produce a partir de la disolución de las partículas de fosfato tricalcico y la precipitación de hidroxiapatita deficiente en calcio de baja cristalinidad, similar a las biológicas, según la reacción:3ALFA-CA3(PO4)2+H2O-CA9(HPO4)(PO4)5(OH) existe una relación lineal directa entre la resistencia mecánica del cemento y el porcentaje de fosfato tricalcico que ha reaccionado. Los estudios microestructurales muestran que la resistencia a la comprensión se puede atribuir al entrelazamiento entre los cristales que precipitan.Existen distintas variables de procesado que afectan de forma significativa a las propiedades de fraguado y endurecimiento del cemento. Entre estas cabe destacar el tamaño de partícula del polvo del cemento, la adición de semillas, la utilización de soluciones de Na2HPO4, la relación liquido/polvo empleada y la temperatura. Tras estudiar el efecto de estas variables sobre algunos parámetros del cemento, como los tiempos de fraguado, el tiempo de cohesión, y la velocidad de endurecimiento se investigan los mecanismos a través de los cuales tienen lugar los efectos / Calcium phosphate cements have attracted much attention in recent years as bone regeneration materials. A calcium phosphate cement consists of a solid phase formed by calcium phosphate powders and a liquid phase, which after mixing are able to set at room or body temperature, producing a solid body Together with the biocompatibility and bioactivity of the calcium phosphate ceramics, calcium phosphate cements have specific advantages derived from their processing route, such as their mouldability, in situ setting ability, which can solve the fixation and adaptation problems inherent to calcium phosphate ceramics.This PhD Thesis is focused on the development and characterisation of a calcium phosphate cement based on the alpha-tricalcium phosphate (alfa-TCP) hydrolysis. The setting of the cement is produced by the dissolution of the alfa-TCP particles and the precipitation of a low crystallinity calcium deficient hydroxyapatite, similar to the biological hydroxyapatite, according to the following reaction: 3ALFA-CA3(PO4)2+H2O-CA9(HPO4)(PO4)5(OH) Both the degree of reaction and the compressive strength increase initially linearly with time, reaching subsequently a saturation level. A direct relation exists between the mechanical strength and the amount of reacted alfa-TCP. The microstructural analysis show that the cement hardening is caused by the entanglement between the precipitated crystals. The reaction mechanisms which control the reaction kinetics in the different stages of reaction are identified based on X-ray diffraction and electron microscopy studies.The contact of the cement with aqueous solutions which simulate body fluids does not affect its properties, and its setting and hardening behaviour at 37ºC fulfil the clinical requirements.Several processing parameters have been identified which affect significantly the setting and hardening properties of the cement. Among them, the particle size distribution of the starting powder, the addition of seed materials, the use Na2HPO4 solutions and the liquid to powder ratio used, are especially relevant. The effect of these parameters on several cement properties, such as the setting and cohesión times, the hardening rate, the maximum strength, the reaction kinetics and the final microstructure is analysed, and the underlying mechanisms are discussed.