Chemical and Biomolecular Engineering - Theses
Permanent URI for this collection
Search Results
1 - 5 of 5
-
ItemApplication of Advanced Fluorescence Imaging Techniques for Intracellular Tracking of Nano-biomaterials( 2021)The engineering and intracellular delivery of nanoparticles with tailored structural, functional and therapeutic properties is challenging due to the interactions of nanomaterials with complex and dynamic biological systems. Additionally, the clinical translation of nanoparticle-based chemotherapeutics is hampered by the poor capacity of 2D cell monolayer culture to mimic in vivo tumour microenvironment and cell-cell interactions. To overcome these biological barriers and enable the clinical translation of nanoparticles, a thorough investigation of nanoparticle-cell interactions in complex biological environments is of paramount importance. For this purpose, novel advanced fluorescence techniques enable the study of nanoparticles structure and functional properties inside the cells with improved spatial and temporal resolution. Additionally, the development of complex 3D cell culture systems mimicking tumour tissue could provide a novel method to predict the in vivo behaviour of nanoparticle-based chemotherapeutics and cellular response to the treatment. Herein DNA- and sugar-based nanoparticles have been developed as platforms for the detection of molecular targets and delivery of drugs within cells and in complex biological settings. Specifically, fluorescence resonance energy transfer microscopy, fluorescence correlation spectroscopy, fluorescence lifetime imaging microscopy and multicolour single-molecule localization microscopy were employed to probe the specific binding of the DNA nanosensor to the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kB) and the nanocomplexation of glycogen with albumin. The intracellular trafficking and the activity of drug-loaded nanoparticles were investigated in 2D and 3D static and dynamic cell culture systems. The biological activity of glycogen-albumin nanoparticles was investigated in a 3D tumour microtissue obtained by co-culturing BT474, NIH-3T3 and RAW264.7 cells in a U-Cup perfusion bioreactor device. The interactions of glycogen-albumin nanoparticles with peripheral blood mononuclear cells isolated from human blood and nanoparticles in vivo biodistribution in mice were also analyzed. This study aims to gain an understanding of the bio-nano interactions in various biological systems and highlights the importance of combining multiple fluorescence techniques and complex models for monitoring the intracellular behaviour of nanomaterials and accurately predicting their in vivo behaviour.
-
ItemFunctional Supramolecular Network Engineering Inspired by Metal–Phenolic Complexation( 2021)Supramolecular assembly provides a versatile pathway for engineering bespoke materials, such as metal–organic hybrid materials. Metal–phenolic networks (MPNs), constructed from the coordination-driven assembly of phenolic ligands and metal ions, are an emerging class of hybrid materials with a rich choice of building blocks. Due to their strong adhesion to different substrates (particles, planar surfaces, microorganisms), high degree of modularity, and tuneable degradability, MPNs have garnered considerable attention in fields such as drug delivery, bioimaging, antimicrobials, separation, and catalysis. However, fundamental research in the material aspects of MPNs and how these influence biomedical applications are essential yet overlooked. This thesis explores the fundamental principles of MPNs and uses this insight to examine MPN materials in a range of biomedical applications. First, MPN microcapsules comprised of various building blocks are engineered, and their programmable gating mechanisms are explored in terms of intermolecular dynamics. This fundamental study not only provides insight into the dynamic nature of MPNs but also offers a route to engineer smart delivery systems and selective gating materials. Next, MPN coatings are used as a versatile and cytocompatible platform to trigger the endosomal escape of nanoparticles, which has been regarded as a key bottleneck for the intracellular delivery of therapeutics. The escape mechanism is systematically investigated and determined to be the “proton-sponge effect”, arising from the buffering capacity of MPNs. Notably, this buffering-enabled escape capability is preserved after the post-functionalization of MPN coatings with polymers, showing the generalizable nature of the platform. Therefore, a subsequent in-depth exploration of the buffering effects of MPNs sheds mechanistic insight into metal–organic systems and their emergent buffering capacity based on coordination dynamics and building block choice. Finally, the advantages of different polyphenol-enabled supramolecular networks are integrated to expand the MPN platform from thin films to self-assembled nanoparticles. Bioactive metal–phenolic nanoparticles are developed via robust and template-free assembly. Hydrophobic interactions and coordination play dominant roles in the assembly and stabilization of the nanoparticles. Furthermore, the incorporation of diverse biomacromolecules (e.g., functional proteins and genes) during assembly enables the potential use of these metal–phenolic nanoparticles in various biomedical applications, anticancer treatments, cascade reactions, and gene knockdown.
-
ItemCryogels for soft tissue engineering and regeneration( 2015)Cell supportive scaffolds that imitate the natural in vivo environment are seen as an essential element for the replacement and regeneration of tissue. Hydrogels have been investigated extensively to this end as they closely mimic the gel-like properties - and in some cases also the chemical properties - of the extracellular matrix (ECM). In particular, macroporous hydrogels containing interconnected macropores (tens to hundreds of microns) have been investigated to support tissue growth as they allow for the diffusion of oxygen and nutrients, and the removal of waste products. More importantly, macroporous structures have also been shown to facilitate the formation of vascular structures. This is most significant as the formation of mature vascular structures plays a key role in the successful integration of large tissue constructs, and remains a key challenge in the field of tissue engineering. Cryogelation, i.e., the formation of hydrogels below the melting temperature of the solvent, is one of many techniques that can be used to fabricate macroporous hydrogels. In this research cryogelation was employed to form macroporous hydrogels from a range of synthetic and naturally based materials in order to explore the potential of cryogels for soft tissue regeneration and tissue engineering. Initially, cryogels were formed from acrylamide, bis-acrylamide and acrylic acid to investigate the cryogelation method in detail. The material and biological properties of the resulting – fully synthetic and non-degradable – gels were assessed using a wide range of characterisation methods. The swelling properties, gel fraction, porosity, pore size and elastic properties of the polyacrylamide cryogels were investigated as a function of precursor composition and reaction conditions. In addition, the biocompatibility of these polyamide cryogels was investigated in vitro using 3T3 fibroblast cells. Collectively these results showed that the cryogelation technique was suitable for forming soft, cell-supportive tissue scaffolds. It thus represents a well-defined and reproducible method to readily afford non-degradable, cell supportive, macroporous ECM mimics. Having shown the suitability of the cryogelation process per se, a modifiable cryogel platform was developed using unmodified hyaluronic acid (HA). HA cryogels were formed at -12 °C, -16 °C and -18 °C and crosslinked with a zero length carbodiimide crosslinker. The physical properties of these gels, including porosity, average pore size, elasticity and swelling were characterised according to the precursor composition and the cryogelation temperature. The resulting cryogels all showed extensive swelling properties and high average porosity (~90%). Cryogels were formed with modifiable pores from 18 ± 2 μm up to 87 ± 5 μm, via manipulations in cryogelation temperature and polymer content. It was possible to readily tune the elastic properties of the cryogels, as measured via rheology, across the entire soft tissue range; ranging from ~ 1 kPa to above 10 kPa. In addition, a method for chemical modification was also explored. Chemical modification of the cryogels importantly improved cell proliferation and cellular interaction, highlighting the potential of the developed platform for biomedical applications. The versatility of the developed hyaluronic acid cryogel platform was subsequently further demonstrated through the fabrication of scaffolds suitable for the in vivo regeneration of adipose tissue and the in vitro replication of the blood stem cell niche as described below. Firstly, cryogels were optimised towards adipose tissue regeneration by incorporating adipose-derived matrix (ADM) into the precursor solution. The resulting macroporous HA-co-ADM hydrogels were shown to drive adipogenesis of 3T3 L1 cells in vitro in the absence of inductive medium and their open porous structure enabled tissue and blood vessel invasion when implanted in vivo without eliciting any noticeable systemic effects or effects on the surrounding tissue. The latter investigation utilised a well-described rat back model and further demonstrated the potential of the HA and ADM gels for applications in soft tissue regeneration. Cryogel scaffolds were implanted in a rat back and assessed after two weeks, during which the HA cryogels scaffolds did not degrade and the open porous structure was maintained. Finally, HA cryogels were developed towards replicating a specific element of the hematopoietic stem cell (HSC) niche; the presentation of immobilised ligands. HA cryogels were functionalised, through a three-step process, with a peptide mimic of the important HSC protein, thrombopoietin. HA scaffolds functionalised with the thrombopoietin mimic, RILL, were investigated in vitro with a factor-dependent cell line, which is unable to survive in the absence of Thrombopoietin (or its mimic). Results from this bioassay indicated that the three-step process was able to functionally couple the RILL peptide to the HA cryogel scaffolds, which in turn supported growth and proliferation of the factor-dependent cells.
-
ItemInvestigations into the mechanical properties and curing characteristics of dental glass-ionomer cements( 2005-11)Conventional glass ionomer cements (GICs), which continue to gain acceptance as superbly biocompatible dental materials, were first released in the early 1970s as a result of research into combining the advantages of silicate cements and polyalkenate cements. The chemistry of GICs is based upon the aqueous reaction between an ion-leachable fluoride glass and polyacid which yields the final cross-linked insoluble ionomer (ionic polymer). The significant advantages of GICs include direct adhesion to tooth structures, fluoride release, minimal dimensional change on curing, significant ease of use and superb biocompatibility, to the extent that affected proximal tooth structures may be retained, remineralised , and strengthened against further caries. GICs have, however, been unfavourably compared with other restorative materials in their mechanical properties and setting characteristics, in particular their relative weakness, the time limitations for the acid-base reaction to proceed to acceptable maturity, and the susceptibility of the immature cement to water sorption or desiccation.
-
ItemNovel biocompatible and biodegradable poly(ethylene glycol)- based scaffolds for soft tissue engineering( 2013)The development of new three-dimensional (3D) biocompatible constructs with improved properties for tissue regeneration is central to advances in the field of tissue engineering. Materials of natural and synthetic origin have been widely investigated for the production of tissue engineering scaffolds via various fabrication methods. Continuing research aims to fabricate scaffolds that possess the important properties of biodegradability, biocompatibility, mechanical integrity, and minimal immunogenicity. Achieving this objective is generally quite difficult since these properties are often dependant on each other and tailoring one property often compromises the other. In this research, we have employed novel synthetic approaches utilising epoxy/amine and acid chloride/alcohol chemistries to prepare poly(ethylene glycol) (PEG)-based scaffolds that are biocompatible and biodegradable, and possess excellent mechanical integrities, making them suitable for various soft-tissue engineering applications. Initially ultrathin Chitosan-PEG hydrogel films (CPHFs) were prepared using epoxy-amine chemistry via diepoxy functionalised PEG, chitosan amines and the co-cross-linker cystamine. The resultant films were very robust and possessed desirable biocompatible and biodegradable properties while supporting the attachment and proliferation of corneal endothelial cells (CECs). To eliminate the issues associated with the use of naturally sourced polymers, we were able to subsequently develop fully synthetic 50 μm thin PEG hydrogel films (PHFs). Acid chloride/alcohol chemistry, together with a facile fabrication method was utilised to produce the PHFs. PHFs were found to possess excellent tensile properties and promoted the in vitro attachment and proliferation of corneal endothelial cells with natural morphologies. In vitro degradation and cytotoxicity studies demonstrated the biodegradable and non-toxic characteristics. In an in vivo ovine model, the hydrogel films adhered naturally onto the interior corneal surface while displaying neither toxicity nor immunogenicity. The PHFs did not hinder the function of the native corneal endothelium, demonstrating their suitability for implantation. To further exploit acid chloride/alcohol chemistry, 3D porous PEG sponges and scaffolds were produced via novel gas foaming, and salt-templating techniques respectively. The rapid exothermic reaction and the HCl gas production results in the formation of highly porous polyester PEG sponges (PPSs), while a salt template was utilised to control pore sizes to produce the hydrogel scaffolds (SPHs). Both PPSs and SPHs possessed excellent mechanical integrities and demonstrated biodegradability and minimal toxicity in vitro. In vivo studies revealed complete infiltration of PPSs and SPHs with vascular tissue within 8 weeks. The porous scaffolds have minimal immunogenicity, and non-toxicity as demonstrated by an in vivo rat model, and can undergo complete degradation to non-toxic products by 16 weeks. The novel PEG hydrogel films and porous scaffolds demonstrate highly desirable physico-chemical properties with excellent biocompatible responses in vivo. Encompassing all the desirable properties of biocompatibility, biodegradability, mechanical integrity and complete tissue integration in vivo, the fabricated scaffolds are excellent candidates for advanced soft-tissue engineering applications.