Thesis (Ph. D.)--University of Wisconsin--Madison, 1957. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 113-117).

Thesis (Ph.D.)--Hong Kong University of Science and Technology, 2006. / Includes bibliographical references (leaves 119-132). Also available in electronic version.

Thesis (M.Phil.)--Hong Kong University of Science and Technology, 2008. / Includes bibliographical references (p. 145-154).

Biomimetic calcium phosphate (CaP) coatings imitate the trabecular bones surface structure and have shown to promote osteogenic differentiation in multipotent cells. The work of this thesis focused on the problem of former CaP coatings cracking and flaking off when being put on a bendable core structure like a 3D-printed poly (ε-caprolactone) (PCL) scaffold. The aim was to provide a chemical linkage between PCL and CaP using a star-shaped polymer (sPEG) and a phosphonate, 2-aminoethylphosphonic acid (2-AEP). First, a published CaP coating protocol was revised and investigated in terms of etching parameters for the PCL scaffold. Results presented reproducible thick coatings for all groups. The protocol was then broadened to include subsequent scaffold incubation in sPEG and 2-AEP solutions. Homogenous CaP coatings of decreased thickness presented themselves, proving feasibility. However, as is often found with physical CaP coating depositions, there were some irregular outcomes even during the same experimental group. A lower consumption of the chemical 2-AEP, for economic reasons, meant that the protocol was altered to simultaneously incubate scaffolds with sPEG and 2-AEP including preceding calculations for molar ratios. For ratios 1:1, 1:2 and 1:3, again a homogenous CaP coating was produced on most of the samples, although reproducibility issues maintained. However, the mechanical bending to induce surface cracking showed that the CaP did strongly bond to the sPEG/2-AEP, while the control CaP coating flaked off the surface in large pieces. This research demonstrates that chemically-bound CaP coatings resist flaking off the fiber surface. Future investigations should focus on the mechanisms of CaP crystallization, to improve reproducibility. / Biomimetische Calciumphosphat (CaP) - Beschichtungen imitieren die oberflächliche Struktur des spongiösen Knochens und wirkten sich bereits begünstigend auf die osteogene Differenzierung von multipotenten Zellen aus. Diese Dissertation konzentriert sich auf das Problem des Reißens und Abplatzens bisheriger CaP-Beschichtungen, wenn diese sich auf einem biegsamen Kern-Gerüst, wie einem 3D-gedruckten Polycaprolacton (PCL)-Konstrukt befanden. Das Ziel war, durch den Gebrauch eines sternförmigen Polymers (sPEG) und eines Phosphonates, 2-Aminoethylphosphonsäure (2-AEP), eine chemische Verknüpfung zwischen PCL und CaP herzustellen. Zuerst wurde ein bereits publiziertes CaP-Beschichtungs-Protokoll nachgestellt und verschiedene Ätzungsparameter untersucht. Die Ergebnisse zeigten reproduzierbare, dicke Beschichtungen in allen Gruppen. Danach wurde dieses Protokoll erweitert, indem es nun nacheinander gestellte Inkubationen in sPEG- und 2-AEP-Lösungen mit einbezog. Dünnere, homogene Beschichtungen waren das Ergebnis, was beweist, dass die Hypothese realisierbar ist. Jedoch zeigten die Ergebnisse nicht reproduzierbare Resultate. Desweiteren, war der 2-AEP Verbrauch nicht wirtschaftlich. Daher wurde das Protokoll weiterentwickelt, indem die Proben, nach vorherigen Berechnungen zu den molaren Verhältnissen, simultan mit sPEG und 2-AEP inkubiert wurden. Für die Verhältnisse 1:1, 1:2 und 1:3 wurden wiederum homogene CaP-Beschichtungen produziert. Mit der Absicht Reproduzierbarkeit zu erzielen, wurden weitere Parameter untersucht. Dies blieb jedoch erfolglos. Zuletzt wurde ein mechanischer Test durchgeführt, welcher eine verbesserte CaP-Adhäsion zu den PCL-Fasern nahelegt, wenn diese zuvor mit sPEG und 2-AEP inkubiert wurden. Zukünftige Untersuchungen werden jedoch von Nöten sein, um Daten zur Oberflächenanalyse und von weiteren mechanischen Tests bereitzustellen und um das Protokoll in Bezug auf die Reproduzierbarkeit zu verbessern.

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Skeletal loss and bone deficiencies are major worldwide problem that is only expected to increase due to increase in aging population. As current standards in treatment autografts and allografts are not without drawbacks, there is a need for alternative bone grafts substitutes. The goal of this project was to utilize electrospinning and heat sintering techniques to create biodegradable full thickness three dimensional biomimetic polymeric scaffolds with macro and nano architecture similar to natural bone for bone tissue engineering. First we have investigated pretreatment with 0.1M NaOH and electrospinning gelatin/PLLA blends as means to increase overall mineral precipitation and distribution throughout the scaffolds when incubated in concentrated simulated body fluid (SBF)10XSBF. Mixture of 10% gelatin and PLLA resulted in the significantly higher degree of mineralization, increased mechanical properties, and scaffolds that supported cellular adhesion and proliferation. In the next step we applied heat sintering technique to fabricate 3D electrospun scaffolds that were used to evaluate effects of mineralization and fiber orientation on scaffold strength. Fiber orientation can make a slight difference in nanofibrous scaffold compressive mechanical properties, but this difference is not as profound as the difference seen with increased mineralization. We also developed a technique to fabricate scaffolds that mimic the organization of an osteon, the structural unit of cortical bone. Resulting scaffolds consisted of concentric layers of electrospun gelatin/PLLA nanofibers wrapped around microfiber core with diameters that ranged from 200-600µm. Individual osteon-like scaffolds were heat sintered to fabricate three dimensional scaffolds contained a system of channels running parallel to the length of the scaffolds, as found naturally in bone tissue. Finally we combined two previously fabricated structures, sintered electrospun sheets and individual osteon-like scaffolds, to create novel scaffolds that mimic dual structural organization of natural bone with cortical and trabecular regions. Mineralization for 24 hr significantly increased mechanical properties of the scaffolds, both yield stress and compressive modulus under physiological conditions. Both nonminerlized and mineralized scaffolds were found to support cellular attachment and proliferation over 28 days in culture, but scaffolds mineralized for 24hr were found to better support osteoblastic differentiation and mineral deposition. / Ph. D.

Calcium phosphate spheres as biomaterials have been attracting attention in recent years. Calcium phosphate occurs naturally in bone, and a hollow structure could be advantageous for drug loading and release. The combination of a calcium phosphate chemistry and a spherical-hollow structure could be an optimal strategy for specific biomaterial applications, e.g., certain dental and drug-delivery applications. The focus of this thesis is on the synthesis, formation mechanism and applications of hollow, spherical calcium phosphate particles. First, the thesis describes two methods for the synthesis of calcium phosphate (CaP) spherical particles. The first method involves synthesis of hollow calcium phosphate spherical particles via a supersaturated buffer solution based on a previous study. It was utilised to prepare spheres for applications in drug delivery and dentistry. The second method was developed to explain the mechanism of formation of hollow calcium phosphate spheres. It aimed at revealing the particular function of magnesium in the formation of spherical particles. With the use of this modified method, it could be concluded that the only ions active in the formation of CaP spherical particles are calcium ions, phosphate ions and magnesium ions. Compared with the thermodynamics of micellisation, a new model, called three ions virtual micelle effect, was developed to explain the mechanism of the Mg function. Following this mechanism, a series of spherical particles of other compositions were explored. These spherical particles included strontium phosphate, barium phosphate, calcium fluoride, strontium fluoride and barium fluoride. In this thesis, CaP spheres were studied for the controlled delivery of active ingredients and as active agent for tooth remineralisation. The first investigated application was to control the release of vancomycin from Poly(methyl methacrylate) (PMMA) cement via strontium-doped CaP spheres (SCPS). The results showed that incorporation of CaP spheres into PMMA could enhance antibiotic release while maintaining the mechanical strength. The second application was to control hydrogen peroxide (HP) release from two bleaching gel, in which CP-loaded CaP spheres were the active ingredient. One gel with low HP concentration was developed as an at-home bleaching gel, and one with high HP concentration was developed as an in-office bleaching gel. The results showed that CaP spheres would give a controlled release of peroxide and thus have a potential to increase the efficacy of the bleaching. The third application was to investigate the potential for an anti-sensitivity effect of the spheres, as active agents in toothpaste. We studied the tooth tubules occlusion and the remineralisation effect of CaP spheres. After 7 days of application, the open dentin tubules and surface were fully covered by a newly formed apatite layer, demonstrating the remineralisation potential of the spheres.

Um dos grandes desafios para o tratamento de defeitos ósseos extensos na região bucomaxilofacial têm sido o desenvolvimento de um biomaterial substituto ósseo ao enxerto autógeno. No presente trabalho avaliou-se a formação óssea e a biodegrabilidade do osso desproteinizado bovino Bio-Oss® e do seu similar GenOx Inorg® e da cerâmica bifásica GenPhos® XP no processo de reparo de defeitos ósseos cranianos em coelhos, comparativamente ao osso autógeno (controle positivo) e coágulo sanguíneo (controle negativo). Foram realizados cirurgicamente defeitos bilaterais de 8-mm de diâmetro nos ossos parietais de 39 coelhos. A seguir os defeitos foram preenchidos aleatoriamente com 0,1cm3 de material ou coágulo conforme cada grupo de tratamento. Após os períodos de 4, 8 e 24 semanas os crânios foram coletados, analisados no microtomógrafo e processados histologicamente. O percentual de volume do defeito ocupado pelo material e osso neoformado foi avaliado pela microtomografia e histomorfometria, enquanto que, para a medula óssea, tegumento e tecido conjuntivo, apenas pela análise histomorfométrica. Os resultados quantitativos obtidos foram comparados estatisticamente pela ANOVA a dois critérios (período e tratamento) e teste de Tukey com p<0,05. A intensidade da associação linear dos dados microtomográficos e histomorfométricos avaliada pelo coeficiente de correlação de Pearson, mostraram correlação moderada a forte. Nos períodos iniciais de reparo (30 e 60 dias), os defeitos tratados com Bio-Oss®, GenOx® Inorg e GenPhos® XP apresentaram manutenção do volume do material enxertado (Vvi médio de 34% ) e formação óssea menor e mais imatura em relação grupo autógeno (Vvi = 22% vs. 32% no grupo autógeno). No período mais tardio (180 dias) a quantidade de formação óssea foi estatisticamente similar nos grupos Bio-Oss® (Vvi = 27%), GenOx® Inorg (Vvi = 26%) e GenPhos® XP (Vvi = 20%) porém, o GenOx® Inorg promoveu a formação de um tecido ósseo mais organizado e com maior acúmulo de biomaterial+osso+medula óssea (Vvi = 67,9%) comparado ao GenPhos® XP (Vvi =58,9%) e Bio Oss (Vvi = 55,6%) mas, inferior ao do enxerto autógeno (Vvi = 78%). Os resultados aqui obtidos permitem concluir que o osso autógeno promove rápida formação e maturação óssea, porém não consegue promover o reestabelecimento completo da díploe removida cirurgicamente. Os materiais BioOss, GenOx® Inorg e GenPhos® XP são excelentes materiais osteocondutores levando a formação óssea em toda extensão do defeito, sendo o GenOx® Inorg o que apresenta menor grau de reabsorção e maior e melhor preenchimento do defeito. / One major challenge for treatment of critical size defects in maxillofacial region has been the development of a substitute biomaterial to the autogenous bone grafts. In present study we evaluated the bone formation and biodegradability of deproteinized bovine bone Bio-Oss® and GenOx® Inorg, and biphasic calcium phosphate GenPhos XP® during bone repair process in rabbits cranial defects compared to autogenous bone (positive control) and blood clot (negative control). In parietal bone of 39 rabbits were made bilateral 8-mm diameter defects, which were filled randomly with 0,1cm3 material or clot as each treatment group. After periods of 4, 8 and 24 weeks skulls of animals were collected, analyzed the MicroCT scanner and histologically processed. The percentage of defect volume occupied by biomaterial and new-formed bone were assessed by histomorphometry and microtomography, while the bone marrow, connective tissue and tegument only by first analysis. The quantitative data were compared by two-way ANOVA analysis (time and treatment) and Tukey\'s test at p <0.05. The intensity of the linear association of MicroCT and morphometric data evaluated by the Pearson correlation coefficient, showed moderate to strong correlation. In the early repair periods (30 and 60 days), the defects treated with Bio- Oss, GenOx® Inorg and GenPhos® XP showed maintenance of the graft material volume (average Vvi of 34%) and lower and more immature bone compared autograft group (Vvi = 22% vs. 32% in the autograft group). In the later period (180 days) the amount of bone formation was statistically similar to the groups Bio-Oss® (Vvi = 27 %), GenOx® Inorg (Vvi = 26%) and GenPhos® XP (Vvi = 20%) however, the bone formation in GenOx® Inorg was more organized and with greater accumulation of particles + bone tissue + bone marrow (Vvi = 67.9%), when compared to GenPhos® XP (Vvi = 58.9%) and Bio-Oss® (Vvi = 55.6%) but lower than the autograft (Vvi = 78%). It was concluded that the autogenous bone promotes rapid bone formation and maturation, but cannot promote the complete reestablishment of diploe surgically removed. The Bio-Oss®, GenOx® Inorg and GenPhos® XP are excellent osteoconductive materials leading to bone formation in the full extent of the defects, and the GenOx® Inorg showing less absorption promotes more and better defect filling.

The purpose of this study was to synthesize calcium phosphates doped with titanium and fluoride ions in different combinations. Pure and doped calcium phosphates were synthesized by a precipitation method. The synthesized materials were sintered at 1100&ordm / C and 1300&ordm / C for 1h. The ceramics were characterized by density measurements to determine the effect of sintering temperature. Presence of phases and bonds were characterized by XRD diffraction and FTIR spectroscopy. Grain sizes of the samples were obtained by SEM. Microhardness test was applied on the samples to determine the mechanical properties of the samples. It was observed that Ti4+ addition decreased the density of samples significantly at 1100&deg / C, whereas increasing the sintering temperature to 1300&deg / C caused an increase. Increasing the F- ion amount increased the densification at 1100&deg / C when molar ratios were 1.0, 1.25, 1.50 and decreased the density at 1300&deg / C when Ca /P molar ratios were 1.0, 1.25, 1.67 and 2.0. Ti4+ and F- co-doped samples showed variety in their density behaviour after the sintering at 1100&ordm / C and 1300&ordm / C. The XRD analyses demonstrated that for Ca to P ratio 1 at 1100&deg / C, &beta / -CPP phase, when sintering temperature was raised to 1300&deg / C, as a second phase of &beta / -CPP and &alpha / -TCP observed. Increasing the molar ratio to 1.25 and 1.50 demonstrated &beta / -TCP and/or &beta / -CPP and &beta / -TCP/ HA at 1100&deg / C and &beta / -TCP and/or &beta / -CPP, &alpha / -TCP, TiO2 and HA, &alpha / -TCP, TiO2 phases at 1300&deg / C, respectively. In higher Ca/P molar ratios of 1.67 and 2.0, HA, &beta / -TCP, &alpha / -TCP, CaO, TiO2, CaTiO3 and HA, CaO, &alpha / -TCP, CaTiO3 phases were determined. Increasing the sintering temperature to 1300&deg / C resulted in transformation to &alpha / -TCP. In FTIR spectroscopy analysis, when the samples with molar ratio of 1, 1.25, 1.50, 1.67 and 2.0, sintered at 1100&deg / C, the characteristic bands of &beta / -CPP, OCP/&beta / -TCP, &beta / -TCP/HA, HA and HA were observed, respectively. With increasing the sintering temperature to 1300&deg / C, samples with molar ratio of 1.0 and 1.25 revealed additional secondary characteristic peaks of HA and &beta / -TCP. SEM images revealed that sintering temperature and ion amounts of dopants had significant effect on grain sizes of the samples. The grain sizes were generally increased when sintering temperature rose from 1100&deg / C to 1300&deg / C. The &mu / -hardness test demonstrated that Ti4+ and F- ions in large amounts had positive effect on the mechanical properties at the sintering temperatures of 1100&deg / C and 1300&deg / C

Anterior cruciate ligament (ACL) rupture is a major clinical problem in sports medicine. The current mainstay of treatment is arthroscopic-assisted ACL reconstruction with a soft tissue tendon graft. However, the affected patients are required to abstain from any pivoting activity for at least six to nine months after the operation to protect the graft-host bone interface in order to allow the graft to heal. In this study, a method to accelerate the graft healing within the bone tunnel is proposed by using a local application of an osteoconductive bone cement (Strontium enriched calcium phosphate cement, Sr-CPC) at the graft-host bone interface. It is postulated that Sr-CPC can induce earlier new bone formation in the gap between the graft and host bone tunnel and hence can result in an accelerated healing of the graft within the bone tunnel in ACL reconstruction. Preparation of Sr-CPCs using the conventional setting method (a dissolution/precipitation process) leads to a delay in setting. This study adopted a new setting reaction, a chelate reaction, to manufacture a Sr-CPC system. The Sr-CPC system was fast-setting, injectable and cohesive, and it was suitable for use in minimally invasive orthopaedics surgeries (e.g. arthroscopic-assisted ACL reconstruction). In order to investigate the biocompatibility and osteoconductivity of the Sr-CPC, in vitro cell experiments and an in vivo animal study were carried out. The in vitro experiments showed that the Sr-CPC was biocompatible with no local toxicity. In addition, a higher proliferation rate of osteoblastic-like MG-63 cells, accompanying higher alkaline phosphatase activity, was found in the Sr-CPC group. The in vivo study using a rat femur metaphyseal bone defect model showed evidence of earlier endochondral ossification which was noted at 2 weeks post operation. Moreover, a higher peri-cement bone formation rate, accompanied by a higher cement resorption rate, was found in the Sr-CPC group at 32 weeks after the operation compared with the convention calcium phosphate cement group. To study the effect of the Sr-CPC on the graft healing within the bone tunnel, a one-stage bilateral ACL reconstruction using an Achilles tendon allograft was performed in 30 rabbits. One study (15 rabbits) was to investigate the effect of the Sr-CPC on the healing of a soft tissue tendon graft within the bone tunnel, and the other study (15 rabbits) was to study the difference between the Sr-CPC and the conventional CPC in the healing of a soft tissue tendon graft within the bone tunnel. The Sr-CPC treated graft showed an accelerated healing at all of the time points when compared with the non-treated graft; and at time points of 3 to 12 weeks when compared with the CPC treated graft. In conclusion, a strontium enriched calcium phosphate cement, which is suitable for the arthroscopic use, was manufactured. It is biocompatible, osteoconductive and degradable. It accelerates the graft healing within the bone tunnels in a rabbit ACL reconstruction model using an Achilles tendon allograft when compared with both of the non-treated group and the conventional CPC-treated group. / published_or_final_version / Orthopaedics and Traumatology / Doctoral / Doctor of Philosophy