An Assessment of Novel Biodegradable Magnesium Alloys for Endovascular Biomaterial Applications

Magnesium alloys have been widely explored as potential biomaterials, but several limitations to using these materials have prevented their widespread use, such as uncontrollable degradation kinetics which alter their mechanical properties. In an attempt to further the applicability of magnesium and its alloys for biomedical purposes, two novel magnesium alloys Mg-Zn-Cu and Mg-Zn-Se were developed with the expectation of improving upon the unfavorable qualities shown by similar magnesium based materials that have previously been explored. The overall performance of these novel magnesium alloys has been assessesed in three distinct phases of research: 1) analysing the mechanical properties of the as-cast magnesium alloys, 2) evaluating the biocompatibility of the as-cast magnesium alloys through the use of in-vitro cellular studies, and 3) profiling the degradation kinetics of the as-cast magnesium alloys through the use of electrochemical potentiodynamic polarization techqnique as well as gravimetric weight-loss methods. As compared to currently available shape memory alloys and degradable as-cast alloys, these experimental alloys possess superior as-cast mechanical properties with elongation at failure values of 12% and 13% for the Mg-Zn-Se and Mg-Zn-Se alloys, respectively. This is substantially higher than other as-cast magnesium alloys that have elongation at failure values that range from 7-10%. Biocompatibility tests revealed that both the Mg-Zn-Se and Mg-Zn-Cu alloys exhibit low cytotoxicity levels which are suitable for biomaterial applications. Gravimetric and electrochemical testing was indicative of the weight loss and initial corrosion behavior of the alloys once immersed within a simulated body fluid. The development of these novel as-cast magnesium alloys provide an advancement to the field of degradable metallic materials, while experimental results indicate their potential as cost-effective medical devices.

Bionano Electronics: Magneto-Electric Nanoparticles for Drug Delivery, Brain Stimulation and Imaging Applications

Nanoparticles are often considered as efficient drug delivery vehicles for precisely dispensing the therapeutic payloads specifically to the diseased sites in the patient’s body, thereby minimizing the toxic side effects of the payloads on the healthy tissue. However, the fundamental physics that underlies the nanoparticles’ intrinsic interaction with the surrounding cells is inadequately elucidated. The ability of the nanoparticles to precisely control the release of its payloads externally (on-demand) without depending on the physiological conditions of the target sites has the potential to enable patient- and disease-specific nanomedicine, also known as Personalized NanoMedicine (PNM). In this dissertation, magneto-electric nanoparticles (MENs) were utilized for the first time to enable important functions, such as (i) field-controlled high-efficacy dissipation-free targeted drug delivery system and on-demand release at the sub-cellular level, (ii) non-invasive energy-efficient stimulation of deep brain tissue at body temperature, and (iii) a high-sensitivity contrasting agent to map the neuronal activity in the brain non-invasively. First, this dissertation specifically focuses on using MENs as energy-efficient and dissipation-free field-controlled nano-vehicle for targeted delivery and on-demand release of a anti-cancer Paclitaxel (Taxol) drug and a anti-HIV AZT 5’-triphosphate (AZTTP) drug from 30-nm MENs (CoFe2O4-BaTiO3) by applying low-energy DC and low-frequency (below 1000 Hz) AC fields to separate the functions of delivery and release, respectively. Second, this dissertation focuses on the use of MENs to non-invasively stimulate the deep brain neuronal activity via application of a low energy and low frequency external magnetic field to activate intrinsic electric dipoles at the cellular level through numerical simulations. Third, this dissertation describes the use of MENs to track the neuronal activities in the brain (non-invasively) using a magnetic resonance and a magnetic nanoparticle imaging by monitoring the changes in the magnetization of the MENs surrounding the neuronal tissue under different states. The potential therapeutic and diagnostic impact of this innovative and novel study is highly significant not only in HIV-AIDS, Cancer, Parkinson’s and Alzheimer’s disease but also in many CNS and other diseases, where the ability to remotely control targeted drug delivery/release, and diagnostics is the key.

Copper ions binding in cu‐alginate gelation

Alginate polysaccharide forms viscous aqueous dispersions and has the ability to form gels in the presence of divalent cations such as calcium and copper. In this work, we have studied cooper ions binding during Cu‐alginate gelation, obtaining quantitative information about the amount and kinetics of cation binding. Our results indicate that copper binding during gelation occurs until a Langmuir‐type equilibrium is reached between bound and free ions in the gel‐contacting solution. The kinetics of metal ions binding can be modeled using Ritchie equation–derived models, allowing the prediction of ionic binding and gel formation temporal evolution. The ratio between cationic and polysaccharide quantities in the gelation system determines the kinetics of gelation and the characteristics of the gel formed. The experimental results and models applied in the work give more insights on alginate gelation and contribute to a reliable design and control of production methods for alginate gel structures.

Kinetic studies of alginate gelation

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PROTEIN-TRIGGERED SUSTAINED DRUG RELEASE CONTROLLED BY APTAMERS BOUND TO OLIGOMER-CROSSLINKED ALGINATE

Sustained drug release systems provide many advantages over traditional delivery methods such as extending the time in which the drug is found to be within an effective concentration within the therapeutic window, which decreases the frequency of administration of the drug, and increases patient compliance. Research using polyacrylamide crosslinked by oligomers containing an aptamer sequence, has demonstrated a pulsatile release over 50 minutes triggered by a 2 mM target adenosine concentration. This thesis aims to build off this concept by designing a system that delivers in a sustained manner when triggered by micromolar target concentrations reflective of disease in vivo, using macromolecular targets. For example, the disease wet age related macular degeneration (wet AMD) is associated with increased concentrations of the protein vascular endothelial growth factor (VEGF-A) – a macromolecule. Patients with wet AMD would benefit from the implantation of devices or microspheres that release drugs in a sustained manner in response to local VEGF concentrations. In this thesis, we hypothesize that the protein lysozyme, used to demonstrate proof-of-concept, could trigger the increased release of drugs from oligomer-crosslinked alginate. The objectives are to (i) demonstrate sustained release from alginate, (ii) design oligomer crosslinked alginate that degrades in response to lysozyme, and then (iii) use these systems to control the release of FITC-dextran with and without lysozyme. A series of control experiments and analyses were used to optimize the crosslinking of alginate by annealed oligomers. The cumulative release of FITC-dextran (MW 20,000) from oligomer crosslinked alginate increased by 3.4 μg when lysozyme (3 μM) was introduced at 48 hours, as opposed to controls which released only 0.2 μg. FITC-loaded alginate microspheres coated by oligomer-crosslinked alginate released 15% more FITC-dextran over 120 hours when placed into 3 μM of lysozyme than without lysozyme. Controls of alginate crosslinked with PEG or control oligomers (without a lysozyme aptamer sequence) had no changes in release with lysozyme. The incorporation of a lysozyme aptamer onto oligomers used to crosslink alginate disks or alginate coatings on microspheres resulted in different diffusion and release of FITC-dextran into PBS with or without lysozyme. This approach could be adapted for the delivery of drugs to diseases with specific protein profiles such as wet AMD.

Engineering design of an integrated sustainable process system for cassava biobased materials

Cassava contributes significantly to biobased material development. Conventional approaches for its bio-derivative-production and application cause significant wastes, tailored material development challenges, with negative environmental impact and application limitations. Transforming cassava into sustainable value-added resources requires redesigning new approaches. Harnessing unexplored material source, and downstream process innovations can mitigate challenges. The ultimate goal proposed an integrated sustainable process system for cassava biomaterial development and potential application. An improved simultaneous release recovery cyanogenesis (SRRC) methodology, incorporating intact bitter cassava, was developed and standardized. Films were formulated, characterised, their mass transport behaviour, simulating real-distribution-chain conditions quantified, and optimised for desirable properties. Integrated process design system, for sustainable waste-elimination and biomaterial development, was developed. Films and bioderivatives for desired MAP, fast-delivery nutraceutical excipients and antifungal active coating applications were demonstrated. SRRC-processed intact bitter cassava produced significantly higher yield safe bio-derivatives than peeled, guaranteeing 16% waste-elimination. Process standardization transformed entire root into higher yield and clarified colour bio-derivatives and efficient material balance at optimal global desirability. Solvent mass through temperature-humidity-stressed films induced structural changes, and influenced water vapour and oxygen permeability. Sevenunit integrated-process design led to cost-effectiveness, energy-efficient and green cassava processing and biomaterials with zero-environment footprints. Desirable optimised bio-derivatives and films demonstrated application in desirable in-package O2/CO2, mouldgrowth inhibition, faster tablet excipient nutraceutical dissolutions and releases, and thymolencapsulated smooth antifungal coatings. Novel material resources, non-root peeling, zero-waste-elimination, and desirable standardised methodology present promising process integration tools for sustainable cassava biobased system development. Emerging design outcomes have potential applications to mitigate cyanide challenges and provide bio-derivative development pathways. Process system leads to zero-waste, with potential to reshape current style one-way processes into circular designs modelled on nature's effective approaches. Indigenous cassava components as natural material reinforcements, and SRRC processing approach has initiated a process with potential wider deployment in broad product research development. This research contributes to scientific knowledge in material science and engineering process design.

Evaluación de desenlace: trasplante meniscal versus segunda meniscectomía

Introducción: El incremento de pacientes sintomáticos de rodilla y la osteoartrosis en jóvenes con limitadas posibilidades terapéuticas después de una meniscectomía, genera la búsqueda de alternativas terapéuticas. A pesar que es poco utilizado en Colombia, el trasplante meniscal es una propuesta para el manejo sintomático. Según cifras norteamericanas, se practican entre 700.000 a 1.500.000 artroscopias de rodilla anualmente, el 50% termina en meniscectomía y de este un 40% persisten sintomáticos. Métodos: Estudio de cohorte retrospectivo, con el objetivo de evaluar dolor (Escala Visual Análoga-EVA) y funcionalidad (Escala de Tegner y Lysholm) en los pacientes a quienes se les realizó trasplante meniscal o meniscectomía por segunda vez, entre los años 2007 a 2015. Resultados: A partir de los 6 meses la EVA mostró una tendencia a la mejoría en el grupo de trasplante meniscal, pasando de Moderado a Leve (p: <0.000). La Escala de Tegner y Lysholm cambió de Pobre a Bueno en el grupo de segunda meniscectomía (p= 0.008) y de Bueno a Excelente en el grupo trasplantado (p=0.225). La calificación promedio de la EVA en el grupo de trasplante presentó mejoría (p=<0.000), a diferencia del grupo de segunda meniscectomía (p=0.591). La escala de Tegner y Lysholm, mostró significancia estadística con tendencia a la mejoría en el grupo de segunda meniscectomía. Discusión: Los resultados muestran que con trasplante meniscal hay mejoría del dolor y la funcionalidad versus un segunda meniscectomía. Para fortalecer la evidencia de este tratamiento son necesarios estudios prospectivos complementarios.

Percutaneous vertebroplasty: a new animal model.

Background Context Percutaneous vertebroplasty (PVP) is a minimally invasive surgical procedure and is frequently performed in humans who need surgical treatment of vertebral fractures. PVP involves cement injection into the vertebral body, thereby providing rapid and significant pain relief. Purpose The testing of novel biomaterials depends on suitable animal models. The aim of this study was to develop a reproducible and safe model of PVP in sheep. Study Design This study used ex vivo and in vivo large animal model study (Merino sheep). Methods Ex vivo vertebroplasty was performed through a bilateral modified parapedicular access in 24 ovine lumbar hemivertebrae, divided into four groups (n=6). Cerament (Bone Support, Lund, Sweden) was the control material. In the experimental group, a novel composite was tested—Spine-Ghost—which consisted of an alpha-calcium sulfate matrix enriched with micrometric particles of mesoporous bioactive glass. All vertebrae were assessed by micro-computed tomography (micro-CT) and underwent mechanical testing. For the in vivo study, 16 sheep were randomly allocated into control and experimental groups (n=8), and underwent PVP using the same bone cements. All vertebrae were assessed postmortem by micro-CT, histology, and reverse transcription-polymerase chain reaction (rt-PCR). This work has been supported by the European Commission under the 7th Framework Programme for collaborative projects (600,000–650,000 USD). Results In the ex vivo model, the average defect volume was 1,275.46±219.29 mm3. Adequate defect filling with cement was observed. No mechanical failure was observed under loads which were higher than physiological. In the in vivo study, cardiorespiratory distress was observed in two animals, and one sheep presented mild neurologic deficits in the hind limbs before recovering. Conclusions The model of PVP is considered suitable for preclinical in vivo studies, mimicking clinical application. All sheep recovered and completed a 6-month implantation period. There was no evidence of cement leakage into the vertebral foramen in the postmortem examination.

To Evaluate the Synergistic Effect of Different Crosslinking Strategies on Collagen Derivatives to Develop Funcional Biomimetic Scaffolds

The preliminary objective of this work was to study how the effect of different crosslinking methodologies can functionally modify various characteristics of biological macromolecules relevant for scaffold development in bone tissue engineering. The research study was classified and studied in three different phases: (i) different crosslinking strategies in gelatin functionalization, (ii) ribose mediated crosslinking in collagen-hydroxyapatite scaffold (iii) different crosslinking mechanisms in functional modification of bone-like scaffold. The obtained results were highly positive in all the three investigated studies. Though the core aim of this research was to explore the available crosslinking strategies in different biological macromolecules, the present study generated significant findings, largely contributing to provide optimum solutions in understanding how the crosslinking density can fine-tune the overall performance of a scaffold, relevant for its functioning in vivo. In particular, this study demonstrated that different crosslinkers at different conditions (pH and temperature) can modify the functional properties of the scaffolds differently, therefore this optimization strategies on these crosslinkers as obtained from this study results will help material scientists in the design and development of bioactive hybrid biomaterials for hard tissue regeneration.

Spatially defined peptidomimetics for pain relief and for targeted cancer therapy and diagnostics

Cancer is a disease that has plagued scientists for decades, and how to treat cancer and its complications are inevitable topics in current scientific research. Cancer pain is a major factor that reduces the quality of life of patients. Therefore, the development of analgesic agents with minimal adverse side effects, especially with low addiction, has attracted more and more attention. Among them, opioid analgesics are widely used to alleviate cancer pain and improve the quality of life of patients with advanced cancer, such as in the palliative therapy. Although peptide drugs are efficient, selective and safe, they have several unignorable disadvantages such as poor biological stability, rapid excretion, difficulty in penetrate blood brain barrier. In order to solve these problems, peptidomimetics were developed by introducing unnatural/modified amino acids, decorated peptide backbone, conformational restrictions and secondary structure mimics in peptide sequence. Compared with peptides, peptidomimetics have improved biological stability, increased bioavailability, high affinity and selectivity for receptor binding, and decreased adverse side effects. As the second part of this thesis, I explored the opportunity to design peptide-functionalized responsive biomaterials for the detection of cancer cell and the selective delivery of cytotoxic drugs. The conjugation of peptides with biomaterials enhanced the stability of the loaded drugs, improved targeted delivery, decreased side effects, and increased bioavailability. The precise and controllable drug delivery platform has profound application prospects in cancer treatment. Grafting specific peptides sequence on the surface of biomaterials can satisfy different drug delivery demands according to the characteristics of both peptides and biomaterials. For example, the introduction of tumor-targeting peptides can guide biomaterials into tumor lesions, and blood-brain barrier (BBB) shuttle peptides can lead biomaterials to penetrate the BBB, etc.

Development and Characterization of Biopolymers

Development and characterization of biopolymers was done in AIJU’s laboratories. AIJU, Technological Institute for children’s products and leisure is based in Spain. The work has the aim to study qualities and characteristics of bioplastics’ blends, in order to design where improvements can be executed. Biopolymers represent a sector with great development possibilities because they combine high technical potential and eco-sustainability. Nowadays, plastic pollution has becoming increasingly concerning, particularly in terms of management of waste. Bioplastics provide an alternative for the disposal of products, reducing the volume of waste and enhancing the end of life recovery. Despite the growing interest in biopolymers there is some gaps that need be filled. The main objective on this work, is the optimization of bioplastics mechanical properties, to find suitable substitutes, as similar as possible to conventional plastics. Firstly, investigations on processability of biomaterials has been deepen since the project deals with toy manufacturing’s sector. Thus, starting from laboratory scale the work aspires to expand industrially. By working with traditional machines, it was notable that, with some limited modifications, the equipment can perform the same functions. Therefore, operational processes do not emerge as an obstacle to the production chain. Secondly, after processing bio-blends, they are characterized by thermal tests (melt flow index, differential scanning calorimetry-DSC, thermogravimetry-TGA) and mechanical tests (traction and flexural tests, Charpy impact, SHORE D hardness and density). While the compatibility does not show relevant results, mechanical improvements has been visualized with addition of more ductile materials. The study was developed by inclusion of sustainable additive VINNEX® to blends. The thesis has highlighted that integration of more flexible materials provides elasticity without compromising bioplastics’ properties.

Study of Bone Biomineralization in 2D and 3D Cultures of a Human Osteosarcoma Cell Line As Osteoblast-like Model

This PhD project focuses on the study of the early stages of bone biomineralization in 2D and 3D cultures of osteoblast-like SaOS-2 osteosarcoma cells, exposed to an osteogenic cocktail. The efficacy of osteogenic treatment was assessed on 2D cell cultures after 7 days. A large calcium minerals production, an overexpression of osteogenic markers and of alkaline phosphatase activity occurred in treated samples. TEM microscopy and cryo-XANES micro-spectroscopy were performed for localizing and characterizing Ca-depositions. These techniques revealed a different localization and chemical composition of Ca-minerals over time and after treatment. Nevertheless, the Mito stress test showed in treated samples a significant increase in maximal respiration levels associated to an upregulation of mitochondrial biogenesis indicative of an ongoing differentiation process. The 3D cell cultures were realized using two different hydrogels: a commercial collagen type I and a mixture of agarose and lactose-modified chitosan (CTL). Both biomaterials showed good biocompatibility with SaOS-2 cells. The gene expression analysis of SaOS-2 cells on collagen scaffolds indicated an osteogenic commitment after treatment. and Alizarin red staining highlighted the presence of Ca-spots in the differentiated samples. In addition, the intracellular magnesium quantification, and the X-ray microscopy on mineral depositions, suggested the incorporation of Mg during the early stages of bone formation process., SaOS-2 cells treated with osteogenic cocktail produced Ca mineral deposits also on CTL/agarose scaffolds, as confirmed by alizarin red staining. Further studies are underway to evaluate the differentiation also at the genetic level. Thanks to the combination of conventional laboratory methods and synchrotron-based techniques, it has been demonstrated that SaOS-2 is a suitable model for the study of biomineralization in vitro. These results have contributed to a deeper knowledge of biomineralization process in osteosarcoma cells and could provide new evidences about a therapeutic strategy acting on the reversibility of tumorigenicity by osteogenic induction.